[go: up one dir, main page]

WO2021016474A1 - Thérapie ibd par inhibition de drd3 dans des lymphocytes t régulateurs - Google Patents

Thérapie ibd par inhibition de drd3 dans des lymphocytes t régulateurs Download PDF

Info

Publication number
WO2021016474A1
WO2021016474A1 PCT/US2020/043313 US2020043313W WO2021016474A1 WO 2021016474 A1 WO2021016474 A1 WO 2021016474A1 US 2020043313 W US2020043313 W US 2020043313W WO 2021016474 A1 WO2021016474 A1 WO 2021016474A1
Authority
WO
WIPO (PCT)
Prior art keywords
treg
mammal
cells
drd3
pharmaceutical composition
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US2020/043313
Other languages
English (en)
Inventor
Rodrigo A. PACHECO RIVERA
Valentina P. UGALDE TAGLE
Francisco J. CONTRERAS CRENOVICH
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Fundacion Ciencia Para La Vida Fcv
Original Assignee
Fundacion Ciencia Para La Vida Fcv
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Fundacion Ciencia Para La Vida Fcv filed Critical Fundacion Ciencia Para La Vida Fcv
Publication of WO2021016474A1 publication Critical patent/WO2021016474A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C12N15/1138—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing against receptors or cell surface proteins
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2320/00—Applications; Uses
    • C12N2320/30—Special therapeutic applications
    • C12N2320/31—Combination therapy

Definitions

  • the present invention provides a method and compositions for suppress and/or inhibiting intestinal inflammation, particularly inflammatory Bowel Diseases (IBD).
  • IBD inflammatory Bowel Diseases
  • the methods and compositions covered for tins invention comprise the therapeutic use of agents that inhibit the biological activity of the dopamine D3 receptor (DRD3) in the domain of IBD.
  • DRD3 dopamine D3 receptor
  • the methods and compositions comprised for this invention arc useful for suppress and/or inhibiting the DRD3 biological activity specifically expressed in CD4+ T-Cells, particularly in the subpopulatian of regulatory CD4 " T-eeUs (Treg), thus attenuating the development of IBD.
  • one of the objects of the present invention is to propose the use of agents that inhibit the biological activity of DRD3 as useful in the preparation of a pharmaceutical composition for the treatment of IBD, particularly Crohn's disease (CD) and ulcerative colitis (UC), and more particularly for UC.
  • the present invention will be an effective treatment for patient with IBD, as for example, CD and UC
  • This invention also comprises therapeutic kits, that comprise DRD3 inhibitors, useful for the IBD treatment, particularly CD and UC.
  • Inflammatory bowel diseases form a group of chronic remittent inflammatory aff ctions of the gastrointestinal tract, among which Crohn's disease (CD) and ulcerative colitis (UC) are the most common.
  • CD Crohn's disease
  • UC ulcerative colitis
  • the overall IBD prevalence approximates 500-900 eases per 100,000 individuals, and has shown a marked increase daring the last decades.
  • Evidence from inflammatory colitis ouse models and from samples obtained from UC and Crohn’s disease (CD) patients has indicated that gut inflammation in IBD is driven mainly by the inflammatory effector CD4 " T-cell subsets T-hdper 1 (Thl ) and I ' M? (Olson ei at., 2011).
  • regulatory €D4 T-eslls (Treg).
  • Treg cells have been shown to be increased in the inflamed lamina intestinal of CD asid UC patients in comparison to nan-inflamed mucosa and mucosa from healthy controls, and l after isolation they retain their ability to suppress effector T-odls in vitro (Maul et al, 2005), thus suggesting that suppressive activity of Treg could be attenuated just in situ b mediators produced by the inflamed gut mucosa.
  • DRD1 -DRD5 DRs
  • DRD1 -DRD5 DRs
  • T celi ⁇ upon inflammation our recen t study showed that /.
  • TNFux TNFux
  • infliximab Remicade
  • adalimumab Humira
  • goHmumab Other biological therapies currently used inhibit the action of the « integral subunit fr aliaoniah, Tysabri), «4(37 integ in (vedolizumab, Entyvio), 1L-I2 and II, -23 preventing binding to its IL-l 2R(fl receptor (ustekkmmah; Stelara),
  • the limitations of the drugs currently used for the treatment of intestinal inflammatory diseases is the non -spec ill city, which inhibits the pathological inflammatory response, but also inhibits the beneficial immune responses that defend us against infectious pathogens and against the development of tumors.
  • TNF- «, 1L- 12 and IL-23 are inflammatory cytokines Invol ved i a general way in the pathogenic inflammatory immune responses, but also in immune responses beneficial tor the organism.
  • integrin-b!ocking antibodies these have a greater degree of specificity since they affect the entry of inflammatory cells into specific tissues.
  • the blockade of «4 by natali mah inhibits the infiltration of lymphocytes both in the central nervous system (blocking a4b! ) and in the intestinal mucosa (blocking a4b7).
  • blockade of a4b7 allows a specific inhibition of tire recruitment of rnllammaiory cells in the intestinal mucosa.
  • this dreg besides inhibiting the entry of inflammatory T cells into the intestinal mucosa, also inhibits the infiltration of regulatory T cells (Tregs) in this tissue, which carry out an immunosuppressive function and therefore beneficial in the context of intestinal inflammatory diseases.
  • Tegs regulatory T cells
  • blockade of 7 edo! smah prevents the entry of T cells into both the colonic mucosa and the mucosa of the small intestine, the sits of elimination of inflammatory cells.
  • the present invention discloses methods and compositions useful for an effective IBD therapy, with high degree of specificity, based on the inhibition the DRD3 biological activity specifically expressed in CD4+ T-Cells. particularly in the subpopu!atioa of Tregs, thus attenuating the development of IBD. Therefore, tbs present invention provides new solutions for the design of improved therapeutic and preventive strategies against IBD, with greater level of effectivity and specificity.
  • the present invention provides therapeutic methods and pharmaceutical compositions useful for the treatment of intestinal inflammation, particularly useful for the treatment of Inflammatory Bowel Diseases (IBD), and more particularly useful for t e treatment of Crohn disease (CD) and ulcerative colitis (UC), and even more particularly useful for ulcerative colitis.
  • IBD Inflammatory Bowel Diseases
  • CD Crohn disease
  • UC ulcerative colitis
  • any of the methods and compositions comprised by this invention may be useful for the effective treatment of intestinal inflammation, particularly for the treatment of IBD, particularly for CD and DC, and even more particularly for UC.
  • the methods and compositions useful for the treatment of IBD comprised by this invention are useful tor blocking or attenuating or inhibiting the biological activity triggered or mediated by DRD3 in regulatory CD4+ T-Cells (Treg),
  • the methods and compositions useful for the treatment of IBD comprised by this invention are useful for blocking or attenuating or inhib ting DRD3-signalling in Treg cells.
  • the methods and compositions useful for the treatment of IBD comprised by this invention which are useful for blocking or atenuating or inhibiting DRD3 -signalling in Treg cells, induce an enhanced Treg suppressive activity.
  • fee methods and compositions useful for fee treatment of 1SD comprised by this invention which are useful for blocking or attenuating or inhibiting DRD3 '-signalling in Treg cells, induce a strongly increase of the Treg got tropism.
  • fee methods and compositions useful for foe treatment of IBD comprised by this invention are for blocking or inhibiting DRD3 -signalling in Treg ceils to induced a potent antiinflammatory effect indaeing an enhanced Treg suppressive activity and a strongly increase of the Treg gut tropism.
  • fee present invention comprises a method for increasing a migration of regulatory T cells into gut mucosa in a mammal, comprising the step of transducing a Treg cell of the mammal with a retroviral vector containing a nucleic acid encoding an U A product that reduces Drd3 transcription in the mammal.
  • the present invention comprises a method for increasing a migration of regulatory T cells into gut mucosa in a mammal, comprising foe ste of transducing a Treg cell of the mammal with a retroviral vector containing a nucleic acid encoding an ENA product that reduces Prd3 transcription in regulatory CD4+ T cells of the mammal.
  • the present invention comprises a method for increasing regulatory T-cells gut tropism, comprising fee step of reduces Drd3 transcription in regulatory CD4T T cells of the mammal Said reduction in the transcription of Drd3 in regulatory CD4 + T cells, although it can be achieved through any of foe biotechnological tools available today ( pharmacological, biological, molecular, chemical, physical, inter alia), preferably in this invention is useful the targeting DRD3- expression on Treg cells, preferably through the generation of retroviral vectors, and even more particularly through viral vectors encoding biomo!ecules (e.g. nucleic acids and/or proteins) able of reducing die transcription ofDrdS.
  • biomo!ecules e.g. nucleic acids and/or proteins
  • foe present invention comprises a method for inducing an enhanced Treg suppressive activity, comprising fee step of reduces rd3 transcription in regulatory CD4+ T cells of the mammal.
  • Said reduction in the transcription of Brd3 in regulatory CD4 + T cells is useful the targeting DRD3- expression on Treg cells, preferably through the generation of retroviral vectors, and even more particularly through viral vectors encoding biomo!eeoles (e.g, nucleic acids and/or proteins) able of reducing the transcription of Drd3.
  • the present invention comprises a method for increasing regulatory T -ceils gut tropism and/or inducing an enhanced Treg suppressive activity, comprising the step of reduces Drd3 transcription in regulatory CD4v T ceils of the mammal.
  • Said reduction in the transcription of Drd3 in regulatory CD4 + T ee!ls although it can be achieved through any of the biotechnological tools available today (pharmacological biological, molecular chemical., physical, inter alia), preferably in this invention is useful the targeting BRD3-expressioo on Treg cells, preferably through die generation of retroviral vectors, and even more particularly through viral vectors encoding biomolecules (e.g. nucleic acids and/or proteins) able of reducing the transcription ofDrcB.
  • biomolecules e.g. nucleic acids and/or proteins
  • the present invention comprises a method fhr increasing regulatory T ⁇ ceils gut tropism and/or inducing an enhanced Treg suppressive activity, comprising the step of blocking or attenuating or inhibiting the biological acti vity triggered or mediated by DRIB in regulatory CB4+ T* Celts (Treg) of a mammal.
  • a method fhr increasing regulatory T ⁇ ceils gut tropism and/or inducing an enhanced Treg suppressive activity comprising the step of blocking or attenuating or inhibiting the biological acti vity triggered or mediated by DRIB in regulatory CB4+ T* Celts (Treg) of a mammal.
  • Said reduction in the biological activity of Dtd3 in regulatory CD4+ T-ceils although it can be achieved through any of the biotechnological tools available today (e.g. pharmacological [e.g. PG01037], biological [e.g.
  • shRNAj preferably in this invention is useful the targeting DRIB -expression on Treg cells, preferably through the generation of retroviral vectors, and even more particularly through viral vectors encoding biomoleeules (e.g. nucleic acids and/or proteins) able of reducing the transcription of 3 ⁇ 4B and/or reducing the biological activity mediated by DRD3.
  • biomoleeules e.g. nucleic acids and/or proteins
  • the present invention comprises a method for attenuating got inflammation in a mammal in need thereof comprising the stop of transducing a Treg coll of the mammal with a retroviral vector containing a nucleic acid encoding an RNA product that reduces Drd3 transcription in the mammal
  • the present invention comprises a method for attenuating gut inflammation in a mammal in need thereof comprising the step of introducing DrdS-defkient Treg cells into the mammal to cause an increase in migration of regulatory T cells into gut mucosa of the mammal aad/or cause an increase in Treg suppressive activity.
  • the present in vention comprises any of the methods and compos? lions mentioned above useful for the treatment of gut inflammation, wherein the agent that reduces I3 ⁇ 4d3 transcription in the mamma! is a nucleic acid encoding an RNA product, where the RNA product is an shRNA or a functionally similar nucleic acid molecule.
  • the present invention comprises any of the methods and compositions mentioned above useful for the treatiuent of gut inflammation, wherein the Drd3-dei ient Treg cells are introduced into the mammal intravenously.
  • this invention comprises methods fur inhibiting intestinal inflammation (e.g. IB ⁇ , CD, UC) in a subject comprising administering to the subject a pharmaceutical composition comprising shRNA and/or any other inhibitor or disrupter agent for blocking or inhibiting DRDS-signailing in Treg cells to induce a potent anti-inflammatory effect * eliciting an enhanced " Frag suppressive activity and a strongly increase of the Treg gut trop ism.
  • die inhibitor or disrupter agent tor blocking or inhibiting DRD3-aigaalling in Treg cells comprised for the pharmaceutical compositions useful for the treatment of intestinal inflammation (e.g. IBIX CD.
  • UC is a chemical molecule or a biological molecule, as for example, peptides, antibodies * nanobodies, aptamers, small ol cule * antagonist, oligonucleotides and any other chemical or biological molecule with de property of blocking or inhibiting DRD3 -signalling in Treg cells to induce a poten anti-inflammatory effect, and in this way eliciting an enhanced Treg suppressive activity and/or a strongly ncrease of the Treg gat fropism.
  • the inhibitor or disrupter agent for blocking or inhibiting Dl 3-signa3img in Treg cells comprised for the pharmaceutical compositions useful for the treatment of intestinal inflammation (e.g. IBD, CD, UC), la a nucleic acid, particularly as R. A, and even more particularly aa shRNA.
  • these nucleic a d are selected from the group of nucleic acid comprised in the Examples of this document or from other sources, which could be normal or modified nucleic acid, where in this last ease, nucleic acid could be modified at terminus and/or Internal regions, with linkers, spacers, special nucleotides, modified bonds, inter alia.
  • the nucleic acids for blocking or inhibiting DRD3- signalling in Treg cells comprised for the pharmaceutical compositions useful for the treatment of intestinal inflammation (e.g, IBD, CD, UC), will generally contain phosphodiesier bonds, although in some eases, oligonucleotides probe analogs are included that may have alternate inter nucleoside linkages, comprising, but not limited to, phosphorothioate, peptide nucleic acid or PNA, phosphoiunnde, phosphorodithioa o.
  • nucleic acids analogs include such as, hut not limited to, morpholine, locked oligonucleotides, peptidic nucleic acids or PNA or 2-o-(2-methoxy) ethyl modified 5 * and 3' end oligonucleotides.
  • the nucleic acids may contain any combination of deoxyribo- and ribonucleotides, and any combination of bases. Including uracil, adenine * thymine, cytosine, guanine, inoslne, xathanine hypoxathanine, isocytosine, isoguanine, inter alia.
  • the methods and compositions useful for the treatment of IBD comprised by this invention which are for blocking or inhibiting RD3 -signalling in Treg cells to induced a potent anti-inflammatory effect, Inducing an enhanced Treg suppressive activity and a strongly Increase of die Treg gut tropism, could be combined with any of the standard therapies actually available for the treatment of IBD.
  • compositions for the treatment of IBD which are for blocking or inhibiting DRD3-signaiiiag in Treg cells to Induced & potent anti-inflammatory effect * inducing an enhanced Treg suppressive activity and a : strongly increase of the Treg gut tropism, where said pharmaceutical composition comprise an of the agents with do property of blocking or inhibiting DRD3-sigrtailmg in Treg cells, and a pharmaceutically acceptable earner * diluent or excipient.
  • the compositions of the present invention can be utilized for therapeutics, diagnostics, prophylaxis and as research reagents an kits. Further, the present invention will be m effective treatment for patient with 18 D, as for example, Crohn Disease and Ulcerative colitis.
  • IB P is as umbrella term used to describe disorders that involve chronic irtfiammation of ie digestive tract.
  • Types of 1BD indude Ulcerative colitis, which is a condition causes long-lasting inflammation and sores (ulcers) in the innermost lining of your large intestine (colon) sod rectum; and Crohn’s disease, which is a type of 18 P that is characterized by inflammation of the lining of your digest? ve tract which often spreads deep into affected tissues. Both ulcerative colitis and Crohn's disease usually involve severe diarrhea * abdominal pais, fatigue and weight loss. 1BD can be debilitating and sometimes leads to life-threatening complications.
  • FIGURES figure L RRD3 ⁇ cMIeienf mice are completely unresponsive to BBS-induced inflammatory solids.
  • Eight-to-ten weeks old Drd3-snfieient (Drd3 ⁇ v* ) and Drd3-deficient (Drd3 ⁇ ) mice were exposed to 1% DSS in the drinking water for 8 d and then switched to normal water.
  • mice were sacrificed and the colon length was determined. Left panel shows representative images of a colon obtained from each experimental group. Right panel shows the quantification. Values arc cm of the whole colon expressed as mean i SEM.
  • CD4' CD45.2' cells obtained from the draining inguinal lymph node were restimulated with FMA and ionomycin in the presence of brefe!din A for 4h and intracellular cytokinemmnnostalmng was performed and analysed by flow cytometry.
  • A Representative dot plots showing the expression oflL-17 and IF -y in the transferred (0045.2 ') a d endogenous (C045.2 ) populations from alive (ZA f) CD4 gated cells, CJD45,2 + producing 1L-17 and IPN-y are framed in the marked ares. Numbers indicate the percentage of CD45.2 ' positive for IL-17 or IFH-y.
  • FIG. 4 Prd3 ⁇ defkient Treg display higher anit-inflammatory effect in BBS-induced inflammatory colitis.
  • A DnfS mRNA transcription in Treg.
  • Naive CD4 ⁇ T-cclls (CD3 CD4 ! CD62L * CD25 “ CD44 ) were isolated from die spleen of wild-type C57BL/6 mice by cell sorting and then incubated under biased conditions to different ate into Treg cells (inducible Treg; iTreg; gray bars) for different times (0, 1 , 2, 3 and 4 d).
  • Treg CD3 ' CD4 €025 K8 ⁇ !
  • Treg results in stronger attenuation of inflammatory colitis Eight- to-ten weeks old wild-type C57BL/6 mice received the i.v. transfer of j 3 ⁇ 43 ⁇ 4£?-suiicient (DrcB '", black symbols) or dd-defieknt (Drd3 ' Q red symbols) Treg cells (CD3 + CD4' CD25 ' cells; 3x 1 (F eells/motse) and then were exposed to 1.75% DSS in the drinking water for 8 d. Afterward, DSS- eontaiaiug drinking water was switched to normal water and mice were monitored for 5 additional days.
  • mice did not receive Treg transfer and was treated only with DSS (white symbols), whilst another group did not receive neither Treg transfer nor DSS (blue symbols).
  • B Body weight was periodically registered throughout the time course of disease development and die percentage of body weight change relati ve to initial weight was quantified.
  • C CD4 " T-cells were isolated from the mesenteric lymph nodes 13 d after Treg transfer and the production of IFN-G and IL- 17 A was determined by intracellular cytokine staining and analysed by flow cytometry. Numbers represent fee percentage of viable (ZAq gate) CD4 " T-cells producing (he corresponding cytokine. (B- €) Data from two independent experiments are shown t> ⁇ 8/group).
  • mice did not receive Treg transfer and was treated only with DSS.
  • Body weight was periodically registered throughout the time course of disease development and the percentage of body weight change relative to initial weight was quantified. Values represeat mean ⁇ SEM Data from four mice per group is shown. Three independent experiments gave similar results *, p ⁇ 0, Q S; comparing Dni3 v Treg ⁇ > DSS versus Drd3 ⁇ Treg -> DSS by two-tailed Student’s t-test.
  • Drd3 ⁇ slgsaiR «g reduces the suppressive activity an fL- ⁇ q prod notion in regulatory T- ceMs in nf
  • naive Drd3 CD45.1 €045,2 CD4 + CD25 T-cells (Tnaive) were loaded with 5 mM CFSE and activated with DCs and anti-CD3 Ab in the presence of CD45.1 CD45.2 ' Treg (at different Tnaive Treg ratios). 72h later, Tnaive proliferation was quantified as the dilution of CFSE- associated fluorescence in the CD45.
  • I CD45.2 CD4 r population by flow cytometry.
  • Tnaive were activated in the absence of Treg (No Tregs).
  • A Tnaive wore eo-eu!tured with freshly isolated r3 ⁇ 4£T v ! or .Drd3"’ Treg.
  • B Prior to the co-culture with Tnaive, Drp ’ Treg were activated with anti-CD3 and anti-CD28 Abs either in the absence or in the presence of the DrdS-seleciive agonist PD 12890? (50 nM) aud incubated for 48h.
  • a and B In left panels representative histograms of CFSE dilution profiles are shown.
  • Markers show the population of naive T cells displaying CFSE dilution. Numbers on the histograms represent the percentage of naive T cells displaying CFSE dilution. In right panels, the extent of suppression is quantified as the percentage of inhibition of naive Teells proliferation relative to maximal proliferation (No Treg), where the percentage of suppression is 0 (dotted line). Dam represent mean ⁇ SEM from three independent experiments. *, p ⁇ 0,0S by impaired Student’s /-test, (C-D) naive Drd3 r: CD45.!
  • Tnaive proliferation was determined as the dilution of CTV-associated fluorescence in the CD45.F €045.2 CD4 : population by flow cytometry,
  • C Representative histograms of CTV dilution profiles are shown. Markers show the population of Tnaive cells displaying CTV dilution. Numbers on the histograms represent the percentage of Tnaive cells displaying CTV dilution.
  • D In right panel , the extent of Tnaive proliferation is quantified. Values represent mean D SEM Data from a representative experiment in triplicate is shown. V p ⁇ 0,0S by enpaired Student’s /-test. nnics., not significant differences were observed.
  • nTreg were isolated from tire mesenteric lymph nodes from Dxd3 ⁇ : Fox S ⁇ or DrdS Foxp3 mice, and then activated with anfi-CD3 and anfi €D28- coated dynabeads (Treg:dynabead ratio ::: 1 :2) in the presence of 0, 100 or 1000 nM dopamine and incubated for 48h. Foxp3 expression was determined by the mean intensity fluorescence (MFI) associated to GPP. Values represent mean D SEM. Data from a representative experiment in triplicate is shown. *, p ⁇ 0.05; **, p ⁇ 0.01 by one-way ANOVA followed by Tukey’s posthoc test.
  • MFI mean intensity fluorescence
  • Trcg cells were restimulated with PMA and ionomycia in the presence of brefeldm A, and intracellular lL-10 was immuaostained and analysed by flow cytometry.
  • XL- 10 production was quantified as the percentage of IL-KT CD4 cells in the ZAq * gate, (6) IL--10 production was determined in the cell culture supernatant by ELISA, (F and G) Values represent mean n SEM. Data from two independent experiments is shown. *, p ⁇ 0.05 by unpaired Student's t-test
  • FIG. 8 DrdS-xignalling attenuates the recruitment of regulatory T-cdls Into the gut mucosa upon inflammation,
  • Treg cells ( €D4 + GFP ) were isolated from mesenteric lymph nodes of Drd3 "; " Foxp. ⁇ !p or Drd3 F xpS ⁇ mice and the expression of CCR9 and «4b7 was analysed in the alive (ZAq ⁇ CD4 + GFP population by flow cytometry. Representative dot plots of CCR9 and «Ab7 expression axe shown in the left panel.
  • Treg cells (CD4 ⁇ GFP : ) were isolated from mesenteric lymph nodes of DrdS * Foxp3 ®P or DrdS '" Faxp3 s!p mice and activated with anti CD3- and anti- €D 8- coated dynabeads (Tregtdynabead ratio ::: 1 :2) in the presence of RA and 1L-2 for 7d.
  • the expression of CCR9 and 4b? was analysed every other day in the alive (2Aqj CD4 ' GFP population by flow cytometry'. Quantification of the frequency and density of CCR9 expression are shown in the top-left and the bottom-left panels respectively.
  • Quantification of the frequency and density of a4b7 expression are shown in the top-middle and fee bottom-middle panels respectively. Quantification of the frequency of cells expressing both together C €R9 and 4b7 is sho n in the top-right panel. Quantification of the density of Foxp3 expression is shown in fee bottom-right panel. Values represent mean ⁇ SEM from triplicates. Data from a representative fro three independent experiments is shown. *, p ⁇ 0 05; **, p ⁇ ( Uli; ***, p ⁇ l.u ' ) ( ll by unpaired Student’s t-test.
  • Treg cells (CD4 + GFP) were isolated from meaenkric lymph nodes t&DrdZ * Foxp3 (Cd45J* * ⁇ and Drd3 " FoxpS ⁇ (Cd43. G : Cd45.2 *1 ⁇ mice,mixed at ratio l ; i and then i.v. transferred ⁇ 10" total eelis per mouse) into wild-type CS7BL/6 (Cd45 *‘ * ) recipient mice 4 d after initiated fee treatment with 1.75% DSS in fee drinking water.
  • mice 24h later, mice were sacrificed and the arrival of Drd3* M (Ci>45 1 * CD45.2 ' ) dlJr S " (CD45.1 * CD4S .2 ) Treg was analysed in the spleen, mesenteric lymph nodes (MLN) colonic lamina intestinal (eLP) and Foyer’s patches by flow cytometry.
  • MN mesenteric lymph nodes
  • eLP colonic lamina intestinal
  • Foyer Foyer’s patches by flow cytometry.
  • the ratio of Drd3 *! -Tmg-to-Z ⁇ £ VFxeg was also analysed before the i.v. adoptive transfer (input).
  • Treg cells (CD4 GFP ’ ) were isolated from mesenteric lymph nodes of Drd3 ' "" Foxp3 or iVriJ" Foxp3 > mice and the expression of Foxp3 (GFP) was analysed in the alive (ZA f) €1)4 " population by flow cytometry . Foxp3 expression was quantified as fee percentage of GFP cells (left panel) or the density of GFP expression (MF1; right panel) in the alive (ZAq ⁇ ) CD4’ gate. Each symbol represent data obtained from a single moose ( :::: 10-14 per group). Lines on the graphs represent mean ⁇ SEM. n.s.. no significant differences were found.
  • Drd3-deficfency results in increased CC 9 expression on ITreg.
  • Naive Cl>4 ' CD62L ' CD44 CD25 T-ceils were isolated if ore Drd3 'f FaxpS ⁇ or Brd ⁇ Foxp mice and activated wife anti ⁇ CO3 and anti-CD28 Ahs and ncubated wife IL-2, RA and TGF-bI for 6 d.
  • Treg were sorted from wiki-type mice, activated for two days and then spinoculated with RV-sfaDRD3 supernatant as in (A). Next, cells were coltared for three additional days with different ratios of DynabeadstTregs. OFF expression was assessed by flow cytometry on fresh cells.
  • a and B Representative density plots are shown. Numbers on framed regions indicate the percentage of OFF " cells among alive (ZAq ) CD4+ cells.
  • Treg cells retain their FoxplF phenotype alter transducing t e with retroviral vectors codifying for shR A for rd3, Treg cells were sorted from wiMfrype mice and then acti vated for 2 d in the presence of IL-2 and anti-CD3/CD28-coate Dynabeads (Dynabeads:Tregs ratio 2:1). Then, cells were washed, spinoculated with RV-shDrd3 supernatant into a retronectin-coated plate and incubated for three additional days with anti-CD3/CD28 ⁇ eoated Dynabeads (ratio DynabeadstTregs ⁇ 2: 1).
  • DAI Disease activity index
  • the present invention which provides methods and compositions useful for an effective IBD therapy, with high degree of specificity, based on the inhibition of the BRD3 biological activity specifically expressed in regulatory €D4 ⁇ T-Cells (Treg), also comprises the following inventive features;
  • a method for treating gut inflammation in a mamma! in need comprising administering to the mammal a pharmaceutical composition comprising one or more agents that inhibit attenuate, disrupt and/or block the biological activity' of DRIB i the mammal.
  • a method for increasing a migration of regulatory T cells into gut mucosa in a mammal in need comprising administering to the mammal a pharmaceutical composition comprising one or more agents that inhibit, attenuate, disrupt and/or block the biological activity of DRD3 in the mammal.
  • a method for increasing regulatory CD4+ T-eells (Treg) suppressive activity into gut mucosa in a mammal in need comprising administering to the mamma! a pharmaceutical composition comprising one or more agents tost inhibit, attenuate, disrupt and/or block the biological activity of DRD3 ia the mam al.
  • any of these methods wherein the one or more agents inhibit, attenuate, disrupt and/or block the biological activity of DRD3 specifically expressed in CD4 ⁇ ; - T-Cells; wherein the one or more agents inhibit, attenuate, disrupt and or block the biological activity of DRD3 specifically expressed in CD4+ T-Cclls, is particularly in the subpopolation of regulatory CD4 - T-ceila (Treg); wherein the one or more agents that inhibit, attenuate, disrupt and/or block the biological activity of DRIB in Treg, reduces rd3 transcription; wherein the one or more agents that inhibit, attenuate, disrupt and/or block the biological activity of DRI>3 in Treg, is an RNA product that reduces ORl>3 transcription; wherein the RNA product is an shRNA; wherein the mammal in need suffer of an inflammatory bowel disease; wherein said inflammatory bowel disease is Crohn Disease; wherein said inflammatory bowel disease is Ulcerative Colitis.
  • a me thod for treating got inflammation in a mamma! in need comprising the s tep of transducing a Treg cell of the mammal with a retroviral vector containing a nucleic acid encoding an RNA product that reduces Drd3 transcription in the mammal,
  • a method for increasing a migration of regulators' T cells into gut mucosa in a mammal in need comprising the step of transducing a Treg ceil of the mammal with a retroviral vector containing a nucleic acid encoding an RNA product that reduces Drd3 transcription in the mammal.
  • a method for increasing Treg suppressive activity into gut mucosa in a mamma! in need comprising the step of transducing a Treg cell of the mammal with a retroviral vector containing a nucleic acid « «coding an .RNA product that reduces Drd3 transcription in the mammal.
  • the R A product is an shRNA; wherein the RNA product have the ability to hybridize under defined conditions to a nucleic acid sequence selected from the group consisting of SEQ ID No.1 (DRD3 mRNA mouse) and SEQ ID Ne.2 (DRIB mRNA human); wherein the RNA product is an shBNA (small hairpin RNA); wherein the shRNA product is defined by SEQ ID No.3.
  • a method for treating gut inflammation in a mammal in need thereof comprising the step of introducing Drd3 ⁇ defieient Treg cells into the mammal to cause an increase in migration of regulatory T cells into gut mucosa of the mammal
  • a method for treating gut inflammation in a mammal in need thereof comprising the step of introducing Drd3-defieie:n! Treg cells into the mammal to cause an increase in Treg suppressive activity into gut mucosa of the mammal.
  • Drd3-def aria Treg cells are introduced into the mammal intravenously; wherein the mammal in need suffer of an inflammatory bowel disease; wherein said inflammatory bowel disease is Crohn Disease; wherein said inflammatory bowel disease is Ulcerative Colitis,
  • the method further comprises administering one or more additional therapies; wherein the one or more additional therapies comprise auli-iuRammatory bowel disease therapy, wherein the additional anti-inflammatory bowel disease therapy comprise therapeutics selected from the group consisting of neutralizers of anti-TNFaipha (infliximab (Remieade), adaimiumab (Humira ⁇ . golimumab (Simponi), Certol unab . alpha integrin s banil inhibitors (naializumab (Tysabrl)), alph b ts?
  • the additional therapies comprise auli-iuRammatory bowel disease therapy
  • the additional anti-inflammatory bowel disease therapy comprise therapeutics selected from the group consisting of neutralizers of anti-TNFaipha (infliximab (Remieade), adaimiumab (Humira ⁇ . golimumab (Simponi), Certol unab . alpha
  • integrin inhibitors (vedol umab (Entyvio)), IL ⁇ 12Rheial blocker that avoid its union to ligands IL-l 2 and JL-23 (ustekinumab (Stelara)), anti-MAd-CAM-l monoclonal antibody (PF-00547659), anti-IL-23 monoclonal antibody (MEDI2070), sphmgosine- 1 -phosphate (SIP) receptor agonist (Ozammod) antisense oligodeoxyimeieotide complementary to mRNA of Smad?
  • SIP sphmgosine- 1 -phosphate
  • Ozammod sphmgosine- 1 -phosphate receptor agonist
  • An agent that inhibit atenuate, disrupt and/or block the biological activity of DRD3 specifically expressed in regulatory CD4+ T-Cells wherein is an RNA preduet that reduces DRD3 transcription; wherein the RNA product have the ability to hybridize wider defined conditions to a nucleic acid sequence selected from the group consisting of SEQ ID No.l (DRD3 mRNA mouse) and SEQ ID o.2 (DRD3 mRNA human); wherein the RNA product is an sliRNA (small hairpin RNA); wherein the shRNA product is defined by SEQ ID Ne.3.
  • a pharmaceutical composition comprising any of die before mentioned agents, and a pharmaceutically acceptable carrier, diluent or excipient; these pharmaceutical compositions, for use in the treatment of inflammatory bowel diseases, particularly for use in the treatment of Crohn disease or Ulcerative Colitis
  • a kit comprising any of the before mentioned pharmaceutical compositions, and instructions for administering the pharmaceutical composition to treat gut inflammation in a mammal, particularly inflammatory bowel disease, and even more particularly for use in the treatment of Crohn Disease or Ulcerative Colitis.
  • kits comprising any of the before mentioned pharmaceutical compositions, and instructions for administering the pharmaceutical composition to increase a migration of Treg into gut mucosa in a mammal in need.
  • a kit comprising any of tire before mentioned pharmaceutical compositions, and instructions for administering the pharmaceutical composition to increase Treg suppressive activity into gut mucosa in a mammal in need.
  • compositions for preparation of a medicament for treatment of inflammatory bowel diseases, particularly for preparation of a medicament tor treatment of Crohn disease or ulcerative colitis.
  • the present invention was developed using the following materials and methods:
  • Wild-type C57BL/6 (Drd “ '* ; C 45.2 ! " ) and Ragf mice were obtained from The Jackson Laboratory.
  • C57BL/6 Dr 3 ⁇ mice were kindly donated by Dr. Mam Caros (Joseph et ai., 20 ( 12).
  • C57BL/6 Ft?xp3 m reporter mice were kindly donated by Alexander Radensky (Fon.ten.ot et al , 2005).
  • Both OT-II and B6.SJL -Ppnf ( Cd45.t ' * ) were kindly provided by Dr, Maria Rosa Bono. Drd OT-H mice, R gf Drd3 "' QMS.
  • Monoclonal antibodies (mAbs) for flow cytometry so.ti-Fos.F3 (clone FJK-16S) conjugated to Phycoeryfhrin (PE)-Cyanine ? (Cy7) and Alloph cocyauin (APC), and aoti-IFN-y (clone XM01.2) conjugated to FE Cy7, anti-adp" (clone BATK32) conjugated to PE and antriCCRP (clone C W.1 .2) conjugated to APC or to AFC ⁇ Cy7 were obtained from sBioseience (San Diego.. CA, USA).
  • Anti-CD4 (clone OKI .5) conjugated to APC and APC ⁇ Cy7; anti-CD25 (done PC61) conjugated to Fluorescein isothiocyanate (FITC); anti-CD44 ( one ⁇ M7) conjugated to PE; anti-CD62L (clone MEL 14) conjugated to APC-Cy7; aoti-fL ⁇ l ?A (clone TC11 181710.1) conjugated to APC; aoti-CD45.2 (clone 104) conjugated to PE-Cy7; anti-CD45 i (clone A20) conjugated to Brilliant Violet (Sv)421; anti- TCRVo.2 (clone B20.1) conjugated to PE aad TCRVPS (clone MR9-4) conjugated to APC were purchased from Biolegend (San Diego, CA, USA).
  • mAbs for Cell Culture the followings Abs low in endotoxins and azide free (LEAF) were purchased from Biolegend.: &nti-CD28 (clone 37,51), aoti-CDBs (clone 145-20 1 ) and suti-IEN-y (clone AN- 18).
  • Carrier-Free cytokines TGF-fl! and 1L-2 were purchased from Bfolegeod.
  • Zombie Aqua (ZAq) Fixable Viability dye detectable by flow cytometry was purchased ftom Biolegend.
  • Phorbol I2 ⁇ myristaie 13-aeetate (PMA), ionomycm ami retinoic acid (RA) were purchased from Sigma- Aldrich (San Lois, MO, USA).
  • Cell Trace Carboxyfiuorescein succinhnidyl ester (CFSE), Brefcldin A and Fetal Bovine Serum (PBS) were obtained from Life Technologies (Carlsbad, C ' A, USA).
  • the peptide derived from the chicken ovalbumin (OVA ⁇ - rw : OT- II peptide or pOT-lI) was purchased from Oenescript (Piseataway, NJ, USA), Freund’s Complete Adjuvant (CFA) and anti-CD3/anti-CD28 conjugated dynabeads were purchased ftom Thermo Scientific, Bovine Serum Albumin (BSA) was purchased to Rockland (Limerick, PA, USA), D$S was obtained Bom MF Biomedicals Drd3 agonist, R ⁇ .128907, was purchased from TOCRIS. Cell trace violet (CTV) was obtained from Invitregen (Carlsbad, CA, USA). Ail tissue culture reagents were bought from Life Technologies.
  • CD4 ⁇ T-eelis were obtained by negative selection of sp!enocytes according to manufacturer Instructions (Miltenyi). Further purification of Treg (CD4 ' CD25 h3 ⁇ 4?5 ) and naive CD4 + T-eells (CD4 ! CD62IZ CD44 CD25 ) ways achieved by labeling enriched CDd* T-celts with the corresponding antibodies and subsequent cell sorting using a FACS Aria II (Bi>), obtaining purities over 98%. Purification of Treg cells from FoxpS** mice was carried out by isolating OFF CD4 * cells by cell sorting.
  • Treg or naive CD4 T-ccl s were st.aioed with 5 mM CFSE or 5 mM CTV and stimulated for 3 d with 50 ng/we0 of plate-bound anti ⁇ CD3 mAh and 2 pg/niL soluble anti-CD28 mAh on fiat-botom 96-well plates (Thermo Scientific). The extent of T-cdi proliferation was determined as the percentage of dilution of CFSE- or CTV-associated fluorescence by flow cytometry.
  • naive CD4 T-cells svete incubated in the conditions indicated above, in the presence of 5 ng/mL TGF-G 1, 10 ug/mL J.L-2 and 100 nM RA. At different incubation times, the cells were assessed for gene and protein expression.
  • cytokine staining analysis cells were restimalated with 1 pg/mL ionomycm and 50 ug/mL PMA for 4 b, in the presence of 5 pg/mL brcfeldin A. Cell surface staining was carried out in PBS with 2% FBS. Fo intracellular staining, cells we e War stained with Zombie Aqua (ZAq) Fixable Viability kit (Biolegend), followed by staining fo cell-surface markers and then resuspended in fixatiou/permeabilization solution (3% BSA and 0.5% saponin in PBS).
  • ZAq Zombie Aqua
  • RNA extracted from cells using the Torn! RNA EZNA kit (Omega Bio-Tck) was DNase-digested using the TURBO DMA-free kit (Ambitm) and 1 gg of RNA was used to synthesize eD A utilizing M-MLV reverse transcriptase, according to manufacturer ⁇ instructions (Life Technologies). Quantitative gene expression analysis was erformed using Brilliant II SYBR Green QPCR Master Mix (Agilent), according to manufacturer's recommendations. Primers were used at a concentration of 0 5 mM. We used 40 PC.R cycles as follows: denatination 30 s at 95° €, annealing 30 s at bCBC and extension 30s at ?27 ' C.
  • Gapdh The sequences of the primers used are the following: BrcB, sense 5 , -GAA €TC €TTAAG € €CCACCAT-3 ! and antisense 5"- GAAGGCCCCGAGCACAAT-3 ! ; and Gapdh, sense S’-TCCGTOTTCCTACCCCCAATG-S’ and atttisense S ⁇ -GAGTGGGAGTTGCTGTTGAAG-Sh fn vitm suppression assays
  • mice were activated with 50 ag of plate-bound anti €D3 and 2 pg/mL soluble anti-CD28 in the absence or presence of 50 nM PD 128907 (TOCR1S).
  • naive T-eclls were loaded with 5 mM CTV and activated with plate-bound anti-CD3 mAh (50 ng) and soluble anii- €D2 ⁇ Ab (2 m-g/mL) and co-euSteed with Treg cither in the absence or in the presence of dopamine (100 :nM or 1000 uM; Sigma -AMrich). After 72h, the extent of naive T-ce!l proliferation was determined as the dilution of CFSE- or CTV- associated fluorescence in the €045.1 " €045.2: 004" population by flow cytometry.
  • WT recipient mice were iv. injected with 3 x 1 " Treg ( €04" €D25 b3 ⁇ 43 ⁇ 4 ) .1 d before starting the administration of 1% or 1 ,75% DSS in the thinking water. BSS was given for a total period of 8 d and then replaced with normal drinking water until the end of the experiment Body weight was recorded throughout the time-course of disease development. The extent of less of the initial body weight as used as the main parameter to determine disease severity. In some experiments, disease activity index (DAI) was also determined as a second readout of disease severity.
  • DAI disease activity index
  • pBullet vector For silencing Drd3 expression, we used the retroviral vector pBullet (Wcijtens et al., 1998). We smelted a region coding ⁇ U6 promoter an shRNA directed to drd3 transcript (5 5 -TOC CCT CTC ITT GGT TTC AAC ACA AC- ) and the Hi promoter, into pBullet vector via Neol and Sail restriction sites (Genscript, Pisco may, NJ). pBullet vector drives the expression of foe entire construct by the CMV promoter upstream the Neol site. This vector was transfected into ftoenix-AMPHO cells.
  • GFF cells were subsequently purified by ceil sorting to generate a stable cell line producing retrovirus coding shRNA for DrdS (RY-shDrd3) in foe supernatant.
  • DrdS DrdS
  • RY-shDrd3 retrovirus coding shRNA for DrdS
  • RY-shDrd3 retrovirus coding shRNA for DrdS
  • RY-shDrd3 retrovirus coding shRNA for DrdS
  • AM values are expressed as the e n ⁇ SEM.
  • Statistical analysis wore performed with two-tailed Student's ( est, when comparing only two groups and with one-way ANOVA followed by Tukey's post-hoc test, when comparing more foan two groups (G phPad Software). P values ⁇ 0,05 were considered significant.
  • DRD3 has been shown to be expressed not only in fee adaptive immune system, but also in neutrophils, eosinophils and Natural Killer cells (McKenna ct al., 2002).
  • Drofhddickncy in naive C04 T-eells results in a selective attenuation of TM -mediated immunity
  • OVA ovalbumin
  • pOTII ovalbumin derived peptide O ' T-II
  • mice receiving the transfer of ilr i-defficient Treg displayed a complete attenuation in the loss of body weight (Fig. 4B).
  • GALT gut-associated lymphoid tissues
  • mice receiving TWJ-deffieieut Treg show a more significant redaction in. Thi 7 frequency than that observed in mice receiving DnS-suffiefetd Treg cells (Fig, 4C), Thus, these results together with those shown in figure 3 and in our previous study (Contreras et ah, 20.16) suggest (hat DR03- signalling in Treg plays a relevant role limiting their suppressive activity preferentially on TM7- mediated responses.
  • Example 3 The selective stimulation of DMB3 attenuates the suppressive activity »f Treg.
  • w first compared the suppressive activity of fh3 ⁇ 4fi-sufficient and i. ⁇ /3-defficient Treg using ⁇ an in vitro suppressive assay. Unexpectedly, we did not find any difference in the suppressive activity in these Treg ceils (Fig, 7A), As mentioned above, Treg cells isolated from inflamed lamina intestinal of IBD patients show impaired suppressive activity, nevertheless after isolation they retain their suppressive activity in vitro (Maui et al., 2005), suggesting that suppressive activity of Treg could be impaired just in situ by mediators produced by the inflamed tissue.
  • Treg The higher attenuation of colitis development exerted by 2 3 deffieient Treg can be due to two non- excluding possibilities: 1 , To a higher suppressive activity, or 2, To an enhanced recruitment of Treg into fee gut mucosa.
  • 1 To a higher suppressive activity
  • 2 To an enhanced recruitment of Treg into fee gut mucosa.
  • MN mesenteric lymph nodes
  • Dr ⁇ B ⁇ defficient mice aad analysed fee expression of CCR9 and «4b?, Importantly, the results show that Ot03 ⁇ d&fieienc) !
  • Treg cells from the MLN of B i-sufficient and iWi-deffieicnt mice and they were activated with anti-CD3/anti-CD28 coated dynabeads in fee presence of RA and 1L ⁇ 2 and the dynamic of CCS9 and a4b7 expression was assessed at different time points during 7 d.
  • Treg cells from the spleen of i>rri5 suffiicient and iWJ-defficient mice and then were activated in vitro in the presence of RA and IL-2 and the dynamic of CCR9 and a4f)7 expression was assessed at different time point during 7 d.
  • the results show that expression of CCR9 and a4b? was similar in DrriJl-sufficient and r3 ⁇ 4s?i-defficient Treg along the time (Fig. IT), suggesting that the regulation of CCR9 expression exerted by DRD3 -signalling seems to be confined to GALT associated Treg cells.
  • I fd-dsficiency increases the acquisition of gut tropism in Treg cells found in the MLN.
  • RV- shDrd3-transdn.eed Treg cells was observed in multiple parameters, including the attenuation in the loss of body weight (Fig, 14A), decreased disease activity index (DAI; Fig. I4B), reduced alterations in the architecture of gut mucosa (Fig, 14C), and a lesser extern of colon shortening (Fig. 14D). Tims, these results show that the reduction of DRD3 expression in Treg cells exerts a potent therapeutic effect dampening gut inflammation.
  • small hairpin RNA (shRNA) against DRD3 in mouse

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
  • Microbiology (AREA)
  • Plant Pathology (AREA)
  • Biophysics (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Physics & Mathematics (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)

Abstract

L'invention concerne une méthode de traitement d'une inflammation intestinale chez un mammifère par administration à celui-ci d'une composition pharmaceutique ayant un ou plusieurs agents qui inhibent, atténuent, perturbent et/ou bloquent l'activité biologique de DRD3 chez le mammifère. L'invention concerne, en outre, un agent qui inhibe, atténue, interrompt et/ou bloque l'activité biologique de DRD3 spécifiquement exprimée dans les lymphocytes T CD4+ régulateurs.
PCT/US2020/043313 2019-07-23 2020-07-23 Thérapie ibd par inhibition de drd3 dans des lymphocytes t régulateurs Ceased WO2021016474A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US201962877720P 2019-07-23 2019-07-23
US62/877,720 2019-07-23

Publications (1)

Publication Number Publication Date
WO2021016474A1 true WO2021016474A1 (fr) 2021-01-28

Family

ID=72322513

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2020/043313 Ceased WO2021016474A1 (fr) 2019-07-23 2020-07-23 Thérapie ibd par inhibition de drd3 dans des lymphocytes t régulateurs

Country Status (2)

Country Link
CL (1) CL2022000179A1 (fr)
WO (1) WO2021016474A1 (fr)

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001089546A2 (fr) * 2000-05-19 2001-11-29 Ancile Pharmaceuticals, Inc. Traitement du syndrome du colon irritable et des troubles associes
WO2008020435A2 (fr) * 2006-08-15 2008-02-21 Quark Pharmaceuticals, Inc Compositions et procédés pour le traitement de troubles de l'humeur
WO2016130845A1 (fr) * 2015-02-11 2016-08-18 Loma Linda University Procédé d'utilisation de cellules dendritiques modifiées pour induire des cellules t régulatrices migrant dans l'intestin et traiter une inflammation intestinale
WO2017168390A2 (fr) * 2016-03-31 2017-10-05 Fundación Ciencia Para La Vida Atténuation de la neurodégénérescence associée à la maladie de parkinson par inhibition du récepteur dopaminergique du type d3 dans les lymphocytes t cd4+

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001089546A2 (fr) * 2000-05-19 2001-11-29 Ancile Pharmaceuticals, Inc. Traitement du syndrome du colon irritable et des troubles associes
WO2008020435A2 (fr) * 2006-08-15 2008-02-21 Quark Pharmaceuticals, Inc Compositions et procédés pour le traitement de troubles de l'humeur
WO2016130845A1 (fr) * 2015-02-11 2016-08-18 Loma Linda University Procédé d'utilisation de cellules dendritiques modifiées pour induire des cellules t régulatrices migrant dans l'intestin et traiter une inflammation intestinale
WO2017168390A2 (fr) * 2016-03-31 2017-10-05 Fundación Ciencia Para La Vida Atténuation de la neurodégénérescence associée à la maladie de parkinson par inhibition du récepteur dopaminergique du type d3 dans les lymphocytes t cd4+

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
FRANCISCO CONTRERAS ET AL: "Dopamine Receptor D3 Signaling on CD4 + T Cells Favors Th1- and Th17-Mediated Immunity", THE JOURNAL OF IMMUNOLOGY, vol. 196, no. 10, 18 April 2016 (2016-04-18), US, pages 4143 - 4149, XP055745254, ISSN: 0022-1767, DOI: 10.4049/jimmunol.1502420 *
RODRIGO PACHECO: "Targeting dopamine receptor D3 signalling in inflammation", ONCOTARGET, vol. 8, no. 5, 31 January 2017 (2017-01-31), pages 7224 - 7225, XP055745262, DOI: 10.18632/oncotarget.14601 *
VIDAL PIA M ET AL: "Targeting the Dopaminergic System in Autoimmunity", JOURNAL OF NEUROIMMUNE PHARMACOLOGY, vol. 15, no. 1, 19 January 2019 (2019-01-19), pages 57 - 73, XP037086463, ISSN: 1557-1890, [retrieved on 20190119], DOI: 10.1007/S11481-019-09834-5 *

Also Published As

Publication number Publication date
CL2022000179A1 (es) 2023-06-02

Similar Documents

Publication Publication Date Title
Méndez-Enríquez et al. Mast cells and their progenitors in allergic asthma
Fang et al. Immune cell dysregulation as a mediator of fibrosis in systemic sclerosis
Jafri et al. Immune regulation of systemic hypertension, pulmonary arterial hypertension, and preeclampsia: shared disease mechanisms and translational opportunities
US10501537B2 (en) Methods for treating cancer
JP2017061551A (ja) 自己免疫関連障害または炎症性障害の治療のための低用量ｉｌ−２の使用
Doki et al. Therapeutic effect of anti-feline TNF-alpha monoclonal antibody for feline infectious peritonitis
JP2019516751A (ja) 同種異系ｔ細胞を用いた自己免疫疾患の処置方法
US20080125390A1 (en) Methods for Modulating Immune and Inflammatory Responses
Chen et al. Progranulin-dependent repair function of regulatory T cells drives bone-fracture healing
CN102552910A (zh) 细胞外基质蛋白1及其调节剂在制备过敏性疾病诊断或治疗药物中的应用
Soleto et al. The BAFF/APRIL axis plays an important role in virus-induced peritoneal responses in rainbow trout
RS61274B1 (sr) Jednolančani oligonukleotidi za upotrebu u medicinskom lečenju poremećaja kože
Yue et al. Dissecting Innate and Adaptive Immunity in Inflammatory Bowel Disease: Immune Compartmentalization, Microbiota Crosstalk, and Emerging Therapies
BR112019014406A2 (pt) métodos de tratar esclerose múltipla usando células t autólogas
US20230272399A1 (en) Inhibitors of line1 and uses thereof
Tosiek et al. CD4+ CD25+ Foxp3+ regulatory T cells are dispensable for controlling CD8+ T cell-mediated lung inflammation
WO2021016474A1 (fr) Thérapie ibd par inhibition de drd3 dans des lymphocytes t régulateurs
Crosson et al. IL-13 promotes sensory-sympathetic neurons crosstalk in asthma
WO2023131323A1 (fr) Nouveaux vaccins à néo-antigènes personnalisés et marqueurs
CN115807090A (zh) Slamf9在肿瘤免疫治疗中的应用
WO2018109540A1 (fr) Méthodes de traitement de maladies associées aux cellules ilc2
EP3635098B1 (fr) Lymphocytes t modifiés pour surexprimer lephf19
Aubert et al. Characterization of a regulatory T cells molecular meta-signature identifies the pro-enkephalin gene as a novel marker in mice
Li et al. Death Receptor 3 Signaling Controls the Balance between Regulatory and Effector Lymphocytes in SAMP1
Dinter et al. PD-1 blockade during T cell priming enhances long-term protection against metastatic tumors by epigenetically tuning T cell exhaustion

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 20765111

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 20765111

Country of ref document: EP

Kind code of ref document: A1