Granted Patent
B2
US 12,661,393 · App. 18/029,599 · Granted Jun 23, 2026
BCG based vaccine compositions and methods of use thereof
Inventors:
Alok Singh (Baltimore, MD); Peter Um (Baltimore, MD); Keira Cohen (Baltimore, MD); William Bishai (Baltimore, MD); Rulin Wang (Baltimore, MD); Srinivasan Yegnasubramanian (Baltimore, MD); Trinity J. Bivalacqua (Philadelphia, PA)
Assignee:
The Johns Hopkins University
View Patent ↗
Loading inventors, assignments & file history…
Abstract
The present disclosure relates to a BCG based therapeutic agent using a BCG strain that overexpresses the STING agonist, c-di-AMP. This BCG strain, called BCG-disA-OE, enhances the elevated trained immunity of macrophages and promotes early anti-viral Type I interferon responses in a subject, providing protection against viral infections such as primary respiratory infections and SARS-CoV-2 infection.
Claims (17)
1 . A method of preventing, ameliorating or treating a viral infection in a subject comprising administering to the subject a recombinant bacille Calmette-Guerin (BCG) bacterial strain of Mycobacterium bovis comprising an exogenous nucleic acid construct comprising:
a mycobacterial promoter and a disA gene,
wherein the disA gene is a Mycobacterium tuberculosis gene, and
wherein the viral infection is a primary respiratory infection thereby treating the infection.
2 . The method of claim 1 , wherein the infection is a SARS-CoV-2 infection.
3 . The method of claim 1 , wherein the subject has a condition selected from the group consisting of obesity, diabetes, cystic fibrosis (CF), non-cystic-fibrosis bronchiectasis, and HIV/AIDS.
4 . The method of claim 1 , wherein the bacterial strain over expresses a stimulator of interferon genes (STING) agonist.
5 . The method of claim 4 , wherein the STING agonist is c-di-AMP.
6 . The method of claim 1 , wherein the disA gene is fused to the mycobacterial promoter.
7 . The method of claim 1 , wherein the nucleic acid construct does not have kanamycin resistance markers.
8 . The method of claim 1 , wherein the bacterial strain is optionally resistant to kanamycin.
9 . The method of claim 1 , wherein the bacterial strain comprises a selectable marker.
10 . The method of claim 1 , wherein the mycobacterial promoter is Hsp60 promoter.
11 . The method of claim 1 , wherein the exogenous nucleic acid construct further comprises a ppiA signal sequence and optionally a spike protein of SARS-COV-2.
12 . The method of claim 1 , wherein the exogenous nucleic acid construct further comprises a ppiA signal sequence and optionally the receptor binding domain of the spike protein of SARS-COV-2.
13 . The method of claim 11 , wherein the nucleic acid construct further comprises a signal peptide.
14 . The method of claim 13 , wherein the signal peptide comprises a ppiA signal sequence.
Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded May 31, 2023
From: WANG, RULIN; YEGNASUBRAMANIAN, SRINIVASAN; BIVALACQUA, TRINITY J.
To: THE JOHNS HOPKINS UNIVERSITY
Reel/Frame 063811/0765 →
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded May 31, 2023
From: SINGH, ALOK; UM, PETER; COHEN, KEIRA; BISHAI, WILLIAM
To: THE JOHNS HOPKINS UNIVERSITY
Reel/Frame 063811/0830 →
Continuity (2)
Provisional Application
63086559
· Oct 1, 2020
References Cited (134)
US 5399363A
· Liversidge et al.
· 1995
[cited by applicant]
US 5466468A
· Schneider et al.
· 1995
[cited by applicant]
US 5543158A
· Gref et al.
· 1996
[cited by applicant]
US 5580579A
· Ruddy et al.
· 1996
[cited by applicant]
US 5629001A
· Michael et al.
· 1997
[cited by applicant]
US 5641515A
· Ramtoola
· 1997
[cited by applicant]
US 5725871A
· Illum
· 1998
[cited by applicant]
US 5756353A
· Debs
· 1998
[cited by applicant]
US 5780045A
· McQuinn et al.
· 1998
[cited by applicant]
US 5792451A
· Sarubbi et al.
· 1998
[cited by applicant]
US 5804212A
· Illum
· 1998
[cited by applicant]
US 6613308B2
· Bartus et al.
· 2003
[cited by applicant]
US 9926346B2
· Fulkerson et al.
· 2018
[cited by applicant]
US 20150071873A1
· Biot et al.
· 2015
[cited by applicant]
CN 106479946A
· 2017
[cited by applicant]
CN 107217026A
· 2017
[cited by applicant]
MX 2015015918A
· 2017
[cited by applicant]
WO WO2008140598A2
· 2008
[cited by applicant]
WO WO2016130616A1
· 2016
[cited by applicant]
WO WO2017011444A1
· 2017
[cited by applicant]
WO WO2017049127A1
· 2017
[cited by applicant]
WO WO2018112360A1
· 2018
[cited by applicant]
WO WO2019203965A1
· 2019
[cited by examiner]
WO WO2020002905A1
· 2020
[cited by applicant]
WO WO2021163602A1
· 2021
[cited by examiner]
CN Office Action in Chinese Application No. 2019800405920, dated Dec. 6, 2023, 15 pages (with English translation).
[cited by applicant]
Fang et al., “ Physiological Function of Second Messenger Cyclic Diguanylate Signaling in
[cited by applicant]
EP Search Report in European Application No. 21754580.5, dated Apr. 10, 2024, 7 pages.
[cited by applicant]
JP Office Action in Japanese Application No. 2020-557965, dated Aug. 2, 2024, 7 pages (with English translation).
[cited by applicant]
Zaczek et al., “Genetic evaluation of relationship between mutations in rpoB and resistance of
[cited by applicant]
Canadian Examination Report for Canadian Patent Application No. 3,097,569 issued Jan. 21, 2025, 5 pages.
[cited by applicant]
Extended European Search Report for European Patent Application No. 21876626.9 issued Feb. 11, 2025, 8 pages.
[cited by applicant]
Rotcheewaphan, S., “Characterization of
[cited by applicant]
Sambandamurthy et al., “A pantothenate auxotroph of
[cited by applicant]
CN Office Action in Chinese Application No. 201980040592 dated May 13, 2023, 17 pages (with English translation).
[cited by applicant]
Rayn, “New therapies in nonmuscle invasive bladder cancer treatment”, Indian J Urol, Mar. 2018, 34(1): 11-19.
[cited by applicant]
Sambandamurthy, “
[cited by applicant]
Ablasser A., et al., “Cell Intrinsic Immunity Spreads to Bystander Cells via The Intercellular Transfer To cGAMP,” Nature, Nov. 28, 2013, vol. 503, No. 7477, pp. 530-534.
[cited by applicant]
Agarwal N., et al., “Cyclic AMP Intoxication Of Macrophages by a
[cited by applicant]
Ahmed D., et al., “Role of Cellular Metabolism in Regulating Type I Interferon Responses: Implications for Tumor Immunology and Treatment,” Cancer Letters, Nov. 28, 2017, vol. 409, pp. 20-29.
[cited by applicant]
Askeland E.J., et al., “Bladder Cancer Immunotherapy: BCG and Beyond,” Advances in Urology, May 2012, vol. 2012, Article ID. 181987, 14 Pages.
[cited by applicant]
Bai Y., et al., “
[cited by applicant]
Bai Y., et al., “Two Dhh Subfamily 1 Proteins In
[cited by applicant]
Barker J.R., et al., “Sting-Dependent Recognition Of Cyclic Di-AMP Mediates Type I Interferon Responses During Chlamydia Trachomatis Infection,” MBio, May-Jun. 2013, vol. 4, No. 3, Article e00018-00013, 11 Pages.
[cited by applicant]
Bharati B.K., et al., “A Full-length Bifunctional Protein Involved In C-di-gmp Turnover Is Required For Long-term Survival Under Nutrient Starvation In
[cited by applicant]
Bowie A.G., et al., “Innate Sensing of Bacterial Cyclic Dinucleotides: More Than Just STING,” Nature Immunology, Dec. 2012, vol. 13, No. 12, pp. 1137-1139.
[cited by applicant]
Burdette D.L., et al., “STING is a Direct Innate Immune Sensor of Cyclic Di-GMP,” Nature, Oct. 27, 2011, vol. 478, pp. 515-518.
[cited by applicant]
Cai X., et al., “The cGAS-cGAMP-STING Pathway of Cytosolic DNA Sensing and Signaling,” Molecular Cell, Apr. 24, 2014, vol. 54, pp. 289-296.
[cited by applicant]
Cho K.H., et al., “
[cited by applicant]
Collins A.C., et al., “Cyclic GMP-AMP Synthase Is an Innate Immune DNA Sensor for
[cited by applicant]
Corrigan R.M., et al., “c-di-AMP Is a New Second Messenger in
[cited by applicant]
Corrigan R.M., et al., “Cyclic di-AMP: Another Second Messenger Enters the Fray,” Nature Reviews Microbiology, Aug. 2013, vol. 11, 29 pages.
[cited by applicant]
Dey B., et al., “A Bacterial Cyclic Dinucleotide Activates The Cytosolic Surveillance Pathway And Mediates Innate Resistance To Tuberculosis,” Nature Medicine, Apr. 2015, vol. 21, No. 4, pp. 401-406, 13 Pages.
[cited by applicant]
Dey B., et al., “Crosstalk between
[cited by applicant]
Dey R.J., et al., “Inhibition of Innate Immune Cytosolic Surveillance by a
[cited by applicant]
Diner E.J., et al., “The Innate Immune Dna Sensor Cgas Produces A Noncanonical Cyclic Dinucleotide That Activates Human STING,” Cell Reports, PMID: 23707065, May 30, 2013, vol. 3, pp. 1355-1361.
[cited by applicant]
Du B., et al., “Functional Analysis of c-di-AMP Phosphodiesterase, GdpP, in
[cited by applicant]
Evans M.R.W., et al., “Squamous Cell Carcinoma Secondary to Buruli Ulcer,” Transactions of the Royal Society of Tropical Medicine and Hygiene, 1999, vol. 93, pp. 63-64.
[cited by applicant]
Extended European Search Report for European Application No. 19787889.5, mailed Jan. 19, 2022, 08 Pages.
[cited by applicant]
Extended European Search Report for European Application No. 19878893.5, mailed Jan. 19, 2022, 09 Pages.
[cited by applicant]
Fitzgerald K.A., et al., “IKKepsilon and TBKI are Essential Components of the IRF3 Signaling Pathway,” Nature Immunology, May 2003, vol. 4, No. 5, pp. 491-496.
[cited by applicant]
Francica B.J., et al., “TNFa and Radioresistant Stromal Cells are Essential for Therapeutic Efficacy of Cyclic Dinucleotide STING Agonists in Nonimmunogenic Tumors,” Cancer Immunology Research, Apr. 2018, vol. 6, No. 4,…
[cited by applicant]
Gao D., et al., “Cyclic GMP-AMP Synthase Is An Innate Immune Sensor Of Hiv And Other Retroviruses,” Science, Aug. 23, 2013, vol. 341, 09 pages.
[cited by applicant]
Gupta K., et al., “Identification, Activity and Disulfide Connectivity of C-di-GMP Regulating Proteins in
[cited by applicant]
Hansen K., et al., “Listeria Monocytogenes Induces Ifni3 Expression Through An I FI 16-, cGAS- And Sting-dependent Pathway,” The EMBO Journal, Aug. 1, 2014, vol. 33, No. 15, pp. 1654-1666.
[cited by applicant]
Hornung V., et al., “OAS proteins and cGAS: Unifying Concepts in Sensing and Responding to Cytosolic Nucleic Acids,” Nature Reviews Immunology, Aug. 2014, vol. 14, No. 8, pp. 521-528.
[cited by applicant]
Hwang J.S., et al., “Synthesis Of Sulfonylurea Conjugated Copolymer Via PEO Spacer And Its In Vitro Short-term Bioactivity In Insulin Secretion From Islets Of Langerhans,” Biomaterials, Jul. 19, 1998, vol. 13, pp. 1189-…
[cited by applicant]
Hwang M.J., et al., “Derivation of Class II Force Fields. VI. Carbohydrate Compounds and Anomeric Effects,” Biopolymers, May 1998, vol. 45 No. 6, pp. 435-468.
[cited by applicant]
Hwang S-J., et al., “Gastric Retentive Drug-Delivery Systems,” Critical Reviews in Therapeutic Drug Carrier System, 1998, vol. 15, No. 3, pp. 243-284.
[cited by applicant]
International Search Report and Written Opinion for International Application No. PCT/US2019/022341, mailed Jul. 4, 2019, 10 Pages.
[cited by applicant]
International Search Report and Written Opinion for International Application No. PCT/US2021/018007, mailed Apr. 29, 2021, 10 Pages.
[cited by applicant]
International Search Report and Written Opinion for International Application No. PCT/US2021/053234, dated Mar. 3, 2022, 12 Pages.
[cited by applicant]
Kalia D., et al., “Nucleotide, c-di-GMP, c-di-AMP, cGMP, CAMP, (p)ppGpp Signaling in Bacteria and Implications in Pathogenesis,” Chemical Society Review, Jan. 7, 2013, vol. 42, No. 1, pp. 305-341.
[cited by applicant]
Kates M., et al., “Intravesical BCG Induces CD4+ T-Cell Expansion in an Immune Competent Model of Bladder Cancer,” Cancer Immunology Research, Jul. 2017, vol. 5, No. 7, pp. 594-603.
[cited by applicant]
Kim M-J., et al., “Caseation Of Human Tuberculosis Granulomas Correlates With Elevated Host Lipid Metabolism,” EMBO Molecular Medicine, May 2010, vol. 2, pp. 258-274.
[cited by applicant]
Konno H., et al., “Cyclic Dinucleotides Trigger ULK1 (ATG1) Phosphorylation of STING to Prevent Sustained Innate Immune Signaling,” Cell, Oct. 24, 2013, vol. 155, No. 3, 24 pages.
[cited by applicant]
Kranzusch P.J., et al., “Structure of Human cGAS Reveals a Conserved Family of Second-messenger Enzymes in Innate Immunity,” Cell Reports, May 30, 2013, vol. 3, No. 5, pp. 1362-1368.
[cited by applicant]
Kubota K., et al., “Identification of 2′-Phosphodiesterase, Which Plays a Role in the 2-5A Systems Regulated by Interferon,” The Journal of Biological Chemistry, Sep. 3, 2004, vol. 279, No. 36, pp. 37832-37841.
[cited by applicant]
Lahaye X., et al., “The Capsids of HIV-1 and HIV-2 Determine Immune Detection of The Viral cDNA By The Innate Sensor Gas In Dendritic Cells,” Immunity, Dec. 12, 2013, vol. 39, No. 6, pp. 1132-1142.
[cited by applicant]
Lamichhane G., et al., “A Postgenomic Method For Predicting Essential Genes At Subsaturation Levels Of Mutagenesis: Application To
[cited by applicant]
Lau L., et al., “DNA Tumor Virus Oncogenes Antagonize the cGAS-STING DNA-sensing Pathway,” Science, Oct. 30, 2015, vol. 350, Issue. 6260, pp. 568-571.
[cited by applicant]
Li L., et al., “Hydrolysis of 2′3′-cGAMP by ENPP1 and Design of Nonhydrolyzable Analogs,” Nature Chemical Biology, Dec. 2014, vol. 10(12), pp. 1043-1048.
[cited by applicant]
Li T., et al., “Human Cytomegalovirus Tegument Protein pUL83 Inhibits IFI16-mediated DNA Sensing for Immune Evasion,” Cell Host & Microbe, Nov. 13, 2013, vol. 14, No. 5, 18 pages.
[cited by applicant]
Li X-D., et al., “Pivotal Roles of cGAS-cGAMP Signaling in Antiviral Defense and Immune Adjuvant Effects,” Science, Sep. 20, 2013, vol. 341(6152), pp. 1390-1394.
[cited by applicant]
Li Y., et al., “Regulating STING in Health and Disease,” Journal of Inflammation, Jun. 2017, vol. 14, No. 11, pp. 1-21.
[cited by applicant]
Liang Q., et al., “Crosstalk Between the cGAS DNA Sensor and Beclin-1 Autophagy Protein Shapes Innate Antimicrobial Mmune Responses,” Cell Host & Microbe, Feb. 12, 2014, vol. 15, No. 2, pp. 228-238.
[cited by applicant]
Malen H., et al., “Definition of Novel Cell Envelope Associated Proteins in Triton X-114 Extracts of
[cited by applicant]
Manikandan K., et al., “Two-step Synthesis And Hydrolysis Of Cyclic Di-AMPIn
[cited by applicant]
Manzanillo P.S., et al., “
[cited by applicant]
Mathiowitz E., et al., “Biologically Erodable Microspheres as Potential Oral Drug Delivery Systems,” Nature, Mar. 27, 1997, vol. 386, pp. 410-414.
[cited by applicant]
Morales A., “BCG: A Throwback from the Stone Age of Vaccines Opened the Path for Bladder Cancer Immunotherapy,” The Canadian Journal of Urology, Jun. 2017, vol. 24, No. 3, pp. 8788-8793.
[cited by applicant]
Office Action for Japanese Patent Application No. 2020557965, mailed Feb. 7, 2023, 12 Pages.
[cited by applicant]
Orzalli M.H., et al., “Nuclear IFI16 Induction of IRF-3 Signaling During Herpesviral Infection And Degradation Of IFI16 by The Viral ICPO Protein,” Proceedings of the National Academy of Sciences of USA, Oct. 30, 2012, …
[cited by applicant]
Parvatiyar K., et al., “The Helicase DDX41 Recognizes The Bacterial Secondary Messengers Cyclic Di-GMP And Cyclic Di-AMP To Activate A Type I Interferon Immune Response,” Nature Immunology, Dec. 2012, vol. 13, pp. 1155-…
[cited by applicant]
Postic G., et al., “Characterization of NmA Homologs From
[cited by applicant]
Powles T., et al., “MPDL3280A (Anti-PD-LI) Treatment Leads to Clinical Activity in Metastatic Bladder Cancer,” Nature, Nov. 27, 2014, vol. 515, No. 7528, pp. 558-562, 12 Pages, XP055207741, DOI:10.1038/nature13904.
[cited by applicant]
Rachman H., et al., “Unique Transcriptome Signature Of
[cited by applicant]
Rao F., et al., “YybT Is A Signaling Protein That Contains A Cyclic Dinucleotide Phosphodiesterase Domain And A GGDEF Domain With ATPase Activity,” Journal of Biological Chemistry, Jan. 1, 2010, vol. 285, No. 1, pp. 473…
[cited by applicant]
Rederlman-Sidi G., et al., “The Mechanism Of Action Of BCG Therapy For Bladder Cancer-a Current Perspective,” Nature Reviews Urology, Mar. 2014, vol. 11, No. 3, pp. 153-162.
[cited by applicant]
Romling U., “Great times for Small Molecules: c-di-AMP, a Second Messenger Candidate in Bacteria and Archaea,” Science Signaling, Aug. 19, 2008, vol. 01, No. 33, 5 Pages.
[cited by applicant]
Rorbach J., et al., “PDE12 Removes Mitocondrial RNA poly(a) Tails and Controls Translation in Human Mitochondria,” Nucleic Acids Research, Jun. 11, 2011, vol. 39, No. 17, pp. 7750-7763.
[cited by applicant]
Schirmer T., et al., “Structural And Mechanistic Determinants Of C-di-Gmp Signalling,” Nature Reviews Microbiology, Oct. 2009, vol. 7, No. 10, pp. 724-735.
[cited by applicant]
Siddiqui M.R., et al., “Current Clinical Trials In Non-muscle Invasive Bladder Cancer,” Urologic Oncology, Aug. 2017, vol. 35, No. 8, pp. 516-527, DOI: 10.1016/j.urolonc.2017.06.043, XP085150983.
[cited by applicant]
Silverman, et aL, Viral Phosphodiesterases That Antagonize Double-stranded to RNase L by Degrading 2-5A, Journal Of Interferon & Cytokine Research, Jun. 2014, vol. 34, No. 6, pp. 455-463.
[cited by applicant]
Srivastav R., et al., “Unique Subunit Packing in Mycobacterial nanoRNase Leads to Alternate Substrate Recognitions in DHH Phosphodiesterases,” Nucleic Acids Research, Jul. 2014, vol. 42, No. 12, pp. 7894-7910.
[cited by applicant]
Sun L., et al., “Cyclic GMP-AMP Synthase Is A Cytosolic DNA Sensor That Activates The Type I Interferon Pathway,” Science, Feb. 15, 2013, vol. 339, No. 6121, pp. 786-791.
[cited by applicant]
Takenaga M., et al., “Microparticle Resins as a Potential Nasal Drug Delivery System for Insulin,” Journal of Controlled Release, 1998, vol. 52, pp. 81-87.
[cited by applicant]
Wang J., et al., “Conservative Change to the Phosphate Moiety of Cyclic Diguanylic Monophosphate Remarkably Affects its Polymorphism and Ability To Bind DGC, PDE, and PilZ Proteins,” Journal of the American Chemical Soc…
[cited by applicant]
Wassermann R., et al., “
[cited by applicant]
Watson R.O., et al., “The Cytosolic Sensor cGAS Detects Mycobacterium tuberculosis DNA to Induce Type I Interferons and Activate Autophagy,” Cell Host and Microbe, Jun. 10, 2015, vol. 17, pp. 811-819.
[cited by applicant]
Witte C.E., et al., “Cyclic Di-AMP is Critical for Listeria Monocytogenes Growth, Cell Wall homeostasis, and Establishment of Infection,” Molecular Biology, May 29, 2013, vol. 4, No. 3, e00282-13, DOI: 10.1128/mBio.0028…
[cited by applicant]
Woodward J.J., et al., “c-di-AMP Secreted by Intracellular Listeria Monocytogenes Activates a Host Type I Interferon Response,” Science, Jun. 25, 2010, vol. 328, No. 5986, pp. 1703-1705.
[cited by applicant]
Wu et al., “Cyclic GMP-AMP is an Endogenous Second Messenger in Innate Immune Signaling by Cytosolic DNA,” Science, Feb. 15, 2013, vol. 339, No. 6121, pp. 826-830.
[cited by applicant]
Wu J.J., et al., “Inhibition of cGAS DNA Sensing by a Herpesvirus Virion Protein,” Cell Host Microbe, Sep. 9, 2015, vol. 18, No. 3, pp. 333-344.
[cited by applicant]
Yang J., et al., “Deletion of the Cyclic Di-AMP Phosphodiesterase Gene (cnpB) in
[cited by applicant]
Zhang Y., et al., “The DNA sensor, Cyclic GMP-AMP Synthase, is Essential for Induction of IFN Beta During Chlamydia trachomatis infection,” The Journal of Immunology, Sep. 1, 2014, vol. 193 No. 5, pp. 2394-2404.
[cited by applicant]
Zhao G.N., et al., “Interferon Regulatory Factors: at the Crossroads of Immunity, Metabolism, and Disease,” Biochimica et Biophysica Acta, Feb. 2015, vol. 1852, No. 2, pp. 365-378.
[cited by applicant]
Zhu D., et al., “Structural Biochemistry of a Vibrio Cholerae Dinucleotide Cyclase Reveals Cyclase Activity Regulation by Folates,” Molecular Cell, Elsevier, Amsterdam, NL, Sep. 18, 2014, vol. 55, No. 6, pp. 931-937, DO…
[cited by applicant]
Zitvogel L., et aL, “Type I Interferons in Anticancer Immunity,” Nature Reviews Immunology, Jul. 2015, vol. 15, pp. 405-414.
[cited by applicant]
Kamat, et al., “BCG Against SARS-CoV-2: Second Youth of an Old Age Vaccine?” Frontiers in Pharmacology, vol. 11, Article 1050, Jul. 9, 2020, 6 pages.
[cited by applicant]
Notice of Reasons for Refusal for JP2023-520398 mailed Aug. 19, 2025, 6 pages.
[cited by applicant]
Tullius, et al., “A Replication-Limited Recombinant
[cited by applicant]
Non-Final Office Action for U.S. Appl. No. 16/638,943 mailed Jun. 13, 2025, 18 pages.
[cited by applicant]