( 12 ) United States Patent

Home , Monobactam, Penam, Ranbezolid

US010391098B2 (12 ) United States Patent (10 ) Patent No. : US 10 , 391, 098 B2 Geller et al. (45 ) Date of Patent: * Aug . 27 , 2019 (54 ) ANTISENSE ANTIBACTERIAL COMPOUNDS 5 ,698 ,685 A 12 / 1997 Summerton et al . 6 , 245 , 747 B1 6 / 2001 Porter et al. AND METHODS 6 , 965, 025 B2 11/ 2005 Gaarde et al. 6 , 969 ,400 B2 11 /2005 Rhee et al. (71 ) Applicants :Oregon State University , Corvallis , 7 ,625 ,873 B2 * 12 /2009 Geller .. . C07F 9 /65583 OR (US ) ; Board of Regents , The 435 /471 University of Texas System , Austin , 7 ,790 ,694 B2 9 /2010 Geller et al. 8 , 067, 571 B2 11 / 2011 Weller et al. TX (US ) 8 , 076 ,476 B2 12 / 2011 Reeves et al. 8 , 299 , 206 B2 10 / 2012 Fox et al. (72 ) Inventors : Bruce L . Geller, Corvallis , OR (US ) ; 8 , 314 ,072 B2 11 / 2012 Geller et al. David Greenberg , Coppell , TX (US ) 8 , 536 , 147 B2 * 9 /2013 Weller .. . . A61K 48/ 00 514 /44 A ( 73 ) Assignees: Board of Regents , The University of 9 , 249 , 243 B2 * 2 / 2016 Weller . . . . A61K 48 / 00 Texas System , Austin , TX (US ) ; 9 ,790 ,495 B2 10 /2017 Geller et al . Oregon State University , Corvallis , 2004 / 0029129 AL 2 / 2004 Wang et al. 2005 / 0288246 AL 12 / 2005 Iversen et al. OR (US ) 2006 / 0241075 Al 10 / 2006 McSwiggen 2006 /0270621 AL 11 /2006 Christiano ( * ) Notice : Subject to any disclaimer , the term of this 2007 /0049542 A1 3 /2007 Geller et al . patent is extended or adjusted under 35 2008 / 0194463 Al 8 / 2008 Weller et al . 2010 / 0016215 A1 1 / 2010 Moulton et al . U . S . C . 154 (b ) by 0 days. 2010 / 0234281 Al 9 / 2010 Weller et al. This patent is subject to a terminal dis 2010 / 0261777 Al 10 / 2010 Shaw et al . 2012 / 0040460 A1 2 / 2012 Rigoutsos et al . claimer . 2012 /0122769 A1 5 / 2012 Iversen 2012 /0213663 A1 8 / 2012 Atieh et al. ( 21 ) Appl . No. : 14 /714 , 104 2012 /0289457 AL 11 /2012 Hanson 2012 /0296087 Al 11 / 2012 Sinha et al . ( 22 ) Filed : May 15 , 2015 2013 /0197220 Al 8 /2013 Veda 2013 /0288369 Al 10 /2013 Iverson 2015 /0141321 A1 5 / 2015 Kole et al . (65 ) Prior Publication Data 2015 /0361425 Al 12 /2015 Geller et al. US 2016 /0106857 A1 Apr. 21 , 2016 FOREIGN PATENT DOCUMENTS Related U . S . Application Data KR 2013 /0005208 A 1 / 2013 ( 60 ) Provisional application No. 62/ 129 ,746 , filed on Mar . WO WO 1993 /001286 A2 1 / 1993 WO WO 2004 / 097017 A2 11 / 2004 6 , 2015 , provisional application No . 62 /000 ,431 , filed Wo WO 2006 / 085973 A2 8 / 2006 on May 19 , 2014 . wo WO 2007/ 009094 A2 1 / 2007 WO WO 2008 / 008113 A1 1 / 2008 (51 ) Int . Cl. WO WO 2009 /005793 A2 1 /2009 A61K 45 /06 ( 2006 .01 ) WO WO 2009/ 064471 A1 5 /2009 C12N 15 / 113 ( 2010 .01 ) ( Continued ) A61K 31 /5377 ( 2006 .01 ) A61K 31 / 712 ( 2006 . 01 ) OTHER PUBLICATIONS A61K 31/ 7036 ( 2006 . 01 ) A61K 47 /64 ( 2017 .01 ) GenBank 01251363 [ KR 1020130005208 - A /39 : Method and kit for ( 52 ) U . S . CI. detecting carbapenem resistant enterobacteriaceae using real - time CPC ...... A61K 31/ 5377 ( 2013 .01 ) ; A61K 31 / 7036 PCR ] ( retrieved on Oct . 13 , 2015 from http : // www . ncbi. nlm .nih . ( 2013 .01 ) ; A61K 31/ 712 (2013 .01 ) ; A61K gov / nucleotide/ 662699238 ? report = genbank & log $ = nuclalign & blast _ 45 / 06 ( 2013 .01 ); A61K 47/ 645 ( 2017 .08 ) ; rank = 1 & RID = 1V403DGD016 ] Jul . 8 , 2014 ( Jul. 8 , 2014 ) whole C12N 15 / 113 ( 2013 .01 ); C12N 2310 / 3233 ( 2013. 01 ) ; C12N 2310 /3513 (2013 .01 ) ; YO2A doc . 50 /473 ( 2018 .01 ) ( Continued ) ( 58 ) Field of Classification Search Primary Examiner — Brian Whiteman CPC ...... A61K 31/ 712 ; C12N 15 / 113; C12N 2310 /3233 ( 74 ) Attorney, Agent, or Firm — Parker Highlander PLLC See application file for complete search history . (57 ) ABSTRACT (56 ) References Cited Provided are antisense oligomers targeted against or genes associated with a biochemical pathway and /or cellular pro U . S . PATENT DOCUMENTS cess , and related compositions and methods of using the 5 ,034 ,506 A 7 / 1991 Summerton et al. oligomers and compositions to treat an infected mammalian 5 , 142 , 047 A 8 / 1992 Summerton et al. subject, for example , as primary antimicrobials or as adjunc 5 , 166 , 315 A 11 / 1992 Summerton et al. tive therapies with classic antimicrobials . 5 , 185 , 444 A 2 / 1993 Summerton et al. 5 ,217 , 866 A 6 / 1993 Summerton et al. 16 Claims, 26 Drawing Sheets 5 , 506 , 337 A 4 / 1996 Summerton et al. 5 , 521 ,063 A 5 / 1996 Summerton et al. Specification includes a Sequence Listing . US 10 ,391 ,098 B2 Page 2

References Cited Summerton et al . 1997 , “ Morpholino antisense oligomers: design , ( 56 ) preparation , and properties. ” Antisense and Nucleic Acid Drug Development ( 1997 ) ; 7 . 3 : 187 - 195 . FOREIGN PATENT DOCUMENTS EP 15795398 .5 , Partial Supplementary European Search Report dated Nov. 29 , 2017 , 7 pages. WO WO 2012 /043730 AL 4 / 2012 Greenberg , et al. , “ Antisense Phosphorodiamidate Morpholino Oligom WO WO 2012 / 064991 A1 5 / 2012 WO WO 2012 / 150960 AL 11 /2012 ers Targeted to an Essential Gene Inhibit Burkholderia cepacia WO WO 2013 /011072 A1 1 / 2013 Complex . " The Journal of Infectious Diseases ( 2010 ) ; 12 : 1822 WO WO 2015 / 175977 A2 11 / 2015 1830 . wo WO 2015 / 179249 AL 11 / 2015 EP 15792493 . 7 , Partial Supplementary European Search Report WO WO 2016 / 108930 A2 7 / 2016 dated Nov . 29 , 2017 , 10 pages. WO WO 2017 / 112885 A16 / 2017 Youngblood , Derek S . , et al. “ Stability of cell -penetrating peptide morpholino oligomer conjugates in human serum and in cells. ” WO WO 2017 / 112888 AL 6 /2017 Bioconjugate Chemistry (2007 ); 18 .1 : 50 -60 . PCT/ US2015 /031150 , International Preliminary Report on Patent OTHER PUBLICATIONS ability dated Nov. 22 , 2016 , 11 pages . PCT/ US2015 /031213 , International Preliminary Report on Patent PCT/ US2015 / 031150 , International Search Report and Written Opin ability dated Nov . 22 , 2016 , 7 pages. ion dated Jan . 14 , 2016 . Nikaido et al. , “ Broad -specificity efflux pumps and their role in PCT/ US2015 /031213 , International Search Report and Written Opin multidrug resistance of Gram -negative bacteria , ” FEMS Microbial ion dated Sep . 2 , 2015 . Rev ., 36 :340 - 363 , 2012 . PCT/ US2015 /000280 , International Search Report and Written Opin ion dated May 2 , 2016 . * cited by examiner U . S . Patent Aug . 27, 2019 Sheet 1 of 26 US 10 , 391, 098 B2

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) ugimL( Tobramycin MIC US 10 , 391, 098 B2 ANTISENSE ANTIBACTERIAL COMPOUNDS ring. Likewise , resistance to aminoglycosides often involves AND METHODS an enzyme capable of inactivating the antibiotic , in this case by adding a phosphoryl, adenyl, or acetyl group . Active CROSS -REFERENCE TO RELATED efflux of antibiotics is another way that many bacteria APPLICATIONS 5 develop resistance . Genes encoding efflux proteins , such as the tetA , tetG , tetL , and tetK genes for tetracycline efflux , This application claims priority under 35 U . S . C . $ 119 ( e ) have been identified . A bacterial target may develop resis to U . S . Application No . 62/ 000 ,431 , filed May 19, 2014 ; and tance by altering the target of the drug . For example, the U . S . Application No . 62 / 129 , 746 , filed Mar. 6 , 2015 ; each of so -called penicillin binding proteins (PBPs ) in many beta which is incorporated by reference in its entirety . 10 lactam resistant bacteria are altered to inhibit the critical antibiotic binding to the target protein . Resistance to tetra STATEMENT REGARDING THE SEQUENCE cycline may involve , in addition to enhanced efflux , the LISTING appearance of cytoplasmic proteins capable of competing with ribosomes for binding to the antibiotic . For those The Sequence Listing associated with this application is 15 antibiotics that act by inhibiting a bacterial enzyme , such as provided in text format in lieu of a paper copy , and is hereby for sulfonamides, point mutations in the target enzyme may incorporated by reference into the specification . The name of confer resistance . the text file containing the Sequence Listing is SATH Escherichia coli normally inhabits the large intestine of 004 _ 01 US _ ST25 . txt. The text file is about 15 KB , was humans as a commensal organism . However, it can also created on May 15 , 2015 , and is being submitted electroni- 20 cause a variety of clinical infections, and is a leading cause cally via EFS -Web . of bacteremia . There has been an alarming increase in the number of antibiotic -resistant strains of E . coli isolated from BACKGROUND patients with nosocomial and community -acquired bacter emia . It is not uncommon for strains to be resistant to Technical Field 25 multiple antibiotics . The present disclosure includes antisense oligomers tar- Acinetobacter baumannii is a ubiquitous organism that geted against bacterial genes involved in a biochemical has emerged over the years to be a significant cause of pathway and/ or cellular process , and related compositions hospital- acquired infections . This change in epidemiology is and methods of using the oligomers and compositions to especially concerning given that A . baumannii has become treat an infected mammalian subject , for example , as pri- 30 one of the most antibiotic - resistant Gram -negative patho mary antimicrobials or as adjunctive therapies with classic gens that the medical community faces world -wide . The antimicrobials . rapid increase in multi -drug resistance in A . baumannii has Description of the Related Art left few therapeutic choices for the treating physician . Drugs Currently , there are several types of antibiotic compounds such as colistin are now frequently used , although colistin in use against bacterial pathogens and these compounds act 35 resistant strains have appeared . Acinetobacter baumannii through a variety of anti -bacterial mechanisms. For can cause a variety of clinical infections, with pneumonia example , beta - lactam antibiotics , such as penicillin and being one of the most frequent. cephalosporin , act to inhibit the final step in peptidoglycan The appearance of antibiotic resistance in many patho synthesis . Glycopeptide antibiotics , including vancomycin genic bacteria , including cases involving multidrug resis and teichoplanin , inhibit both transglycosylation and trans - 40 tance (MDR ), raises the fear of a post -antibiotic era in which peptidation of muramyl- pentapeptide , again interfering with many bacterial pathogens were simply untreatable by medi peptidoglycan synthesis . Other well -known antibiotics cal intervention . Thus, there is a need for antimicrobial include the quinolones , which inhibit bacterial DNA repli - agents that ( i ) are not subject to the principal types of cation , inhibitors of bacterial RNA polymerase , such as antibiotic resistance currently hampering antibiotic treat rifampin , and inhibitors of enzymes in the pathway for 45 ment of bacterial infection , ( ii ) can be developed rapidly and production of tetrahydrofolate , including the sulfonamides . with some reasonable degree of predictability as to target Some classes of antibiotics act at the level of protein bacteria specificity , (iii ) are effective at low doses, and ( iv ) synthesis . Notable among these are the aminoglycosides , show few side effects . such as kanamycin and gentamicin . This class of compounds targets the bacterial 30S ribosome subunit , preventing the 50 BRIEF SUMMARY association with the 50S subunit to form functional ribo somes . Tetracyclines , another important class of antibiotics , Embodiments of the present disclosure relate , in part, to also target the 30S ribosome subunit , acting by preventing the discovery that the antisense targeting of bacterial genes alignment of aminoacylated tRNA ' s with the corresponding associated with biochemical pathways, cellular processes , mRNA codon . Macrolides and lincosamides , another class 55 and / or antibiotic resistance can increase the antibiotic sus of antibiotics, inhibit bacterial synthesis by binding to the ceptibility of otherwise antibiotic - resistant pathogenic bac 50S ribosome subunit, and inhibiting peptide elongation or teria , and reduce the ability of certain pathogenic bacteria to preventing ribosome translocation . grow . For example , the antisense targeting of genes associ Despite impressive successes in controlling or eliminating ated with RNA biosynthesis , protein biosynthesis , fatty acid bacterial infections by antibiotics , the widespread use of 60 biosynthesis , peptidoglycan biosynthesis , cellular energy antibiotics both in human medicine and as a feed supplement homeostasis , cell division , aromatic compound biosynthesis , in poultry and livestock production has led to drug resistance and antibiotic resistance was shown to increase the cell in many pathogenic bacteria. Antibiotic resistance mecha killing and /or antibiotic susceptibility of antibiotic resistant nisms can take a variety of forms. One of the major (e . g ., multi - drug resistant ) bacteria to many commonly used mechanisms of resistance to beta lactams, particularly in 65 antibiotics . In many instances , the antisense oligomers Gram - negative bacteria , is the enzyme beta - lactamase , described herein were shown to be bactericidal at clinically which renders the antibiotic inactive by cleaving the lactam relevant concentrations and to display synergy with multiple US 10 , 391 , 098 B2 classic antibiotics in Acinetobacter and Escherichia , includ where each R4 is independently C . -C . alkyl, and R is ing multiple drug -resistant (MDR ) strains. The antisense selected from an electron pair and H , and Róis selected from oligomers described herein could thus find utility in the OH , - N (R7 ) CH C ( O )NH2 , and a moiety of the formula : treatment of such bacteria , for instance, in combination with antibiotics or as standalone therapies. Embodiments of the present disclosure therefore include a substantially uncharged antisense morpholino oligomer, N - R8, composed of morpholino subunits and phosphorus - contain ing intersubunit linkages joining a morpholino nitrogen of one subunit to a 5' - exocyclic carbon of an adjacent subunit , where : and having ( a ) about 10 - 40 nucleotide bases , and ( b ) a R ? is selected from H and C . - C . alkyl, and targeting sequence of sufficient length and complementarity R8 is selected from G , - C ( O ) - R ' OH , acyl, trityl , and to specifically hybridize to a bacterial mRNA target 4 -methoxytrityl , where : sequence that encodes a protein associated with a biochemi- 1615 R® is of the formula ( O - alkyl) , - wherein y is an cal pathway and / or cellular process, or a protein associated integer from 3 to 10 and each of with antibiotic resistance , as described herein . In some the y alkyl groups is independently selected from C2- C6 instances , the oligomer is conjugated to a cell - penetrating alkyl; peptide (CPP ) . each instance of R ' is N (R19 ) R11 wherein each R1° is In certain embodiments , the targeting sequence is selected 20 independently C , -C6 alkyl, and Ril is from Tables 2A - B . In some embodiments , the oligomer is selected from an electron pair and H ; about 10 - 15 or about 11 - 12 nucleotide bases in length and has a targeting sequence selected from Tables 2A - B . R² is selected from H , G , acyl, trityl, 4 -methoxytrityl , In certain embodiments , an antisense oligomer of the benzoyl, stearoyl, and a moiety of the formula : disclosure is of formula ( I ) : 25

2 Nu

PRI N

( R 12 2N ' N N ( R122, 0 Nu where L is selected from _ C ( O ) (CH2 ) . C ( 0 ) and 40 C ( O ) (CH2 ) 2S2 (CH2 ) 2C ( O ) - , and each R12 is of the for mula (CH , ) , OC ( O ) N (R14 ) , wherein each R14 is of the o = P - R formula ( CH2) 6NHCE = NH )NH2 ; and R is selected from an electron pair , H , and C , - C . alkyl , wherein G is a cell penetrating peptide (“ CPP ” ) and linker 45 moiety selected from - Nu C ( O )( CH, ANH- CPP, C ( O )( CH2 )- NH- CPP, C ( O ) ( CH3) 2NHC ( O )( CH ) NH- CPP, and C ( O )CH NH -CPP , or G is of the formula : 50 R2 R3 CPP , or a pharmaceutically acceptable salt thereof, where each Nu is a nucleobase which taken together forms a targeting sequence ; 55 X is an integer from 9 to 38 ; T is selected from OH and a moiety of the formula : wherein the CPP is attached to the linker moiety by an amide bond at the CPP carboxy terminus , with the proviso that only - R6 60 one instance of G is present, wherein the targeting sequence specifically hybridizes to 0 = — N (R4 ) 2R5 , a bacterial mRNA target sequence that encodes a protein associated with a biochemical pathway and / or cellular pro cess or a protein associated with antibiotic resistance , as 65 described herein . In some embodiments , the target sequence comprises a translational start codon of the bacterial mRNA and /or a US 10 , 391 , 098 B2 sequence within about 30 bases upstream or downstream of comprises a variant having at least 80 % sequence identity to the translational start codon of the bacterial mRNA . SEQ ID NOS: 12 -53 , where thymine bases ( T ) are option In some embodiments , the protein associated with a ally uracil bases (U ) . biochemical pathway and / or cellular process is a fatty acid In certain embodiments , the protein associated with anti biosynthesis protein . In certain embodiments, the fatty acid 5 biotic resistance is selected from at least one of TEM biosynthesis protein is an acyl carrier protein . In certain beta - lactamase (BlaT ), chloramphenicol resistance gene embodiments , the acyl carrier protein is AcpP . In some Cml, and resistance -nodulation - cell division (RND ) - type embodiments , the target sequence is SEQ ID NO : 69 or 70 , and where thymine bases ( T ) are optionally uracil bases ( U ) . multidrug efflux pump subunit AdeA ( adeA ) . In particular In certain embodiments , the fatty acid biosynthesis protein 10 embodiments , the targeting sequence is set forth in SEQ ID is a carboxyltransferase alpha subunit of an acetyl Coen NOS: 54 - 56 , comprises a fragment of at least 10 contiguous zyme A carboxylase . In certain embodiments , the carboxyl nucleotides of SEQ ID NOS : 54 - 56 , or comprises a variant transferase alpha subunit of an acetyl Coenzyme A carboxy having at least 80 % sequence identity to SEQ ID NOS : lase is AccA . In certain embodiments , the targeting sequence 54 -56 , and where thymine bases ( T ) are optionally uracil is set forth in SEQ ID NOS : 1 -11 , comprises a fragment of 15 bas at least 10 contiguous nucleotides of SEQ ID NOS: 1 - 11, or Also included are pharmaceutical compositions, compris comprises a variant having at least 80 % sequence identity to ing a pharmaceutically acceptable carrier and an antisense SEQ ID NOS : 1 - 11, where thymine bases ( T ) are optionally oligomer described herein . Some pharmaceutical composi uracil bases ( U ) . tions further comprising an antimicrobial agent as described In some embodiments , the protein associated with a 20 herein , such as one or more of tobramycin , meropenem , biochemical pathway and / or cellular process is a peptido - and / or colistin . glycan biosynthesis protein . In certain embodiments , the Some embodiments include methods of reducing expres peptidoglycan biosynthesis protein is a UDP - N -acetylglu - sion and activity of a protein selected from at least at least cosamine 1 -carboxyvinyltransferase . In particular embodi- one of a protein associated with a biochemical pathway ments , the UDP - N - acetylglucosamine 1 -carboxyvinyltrans - 25 and /or cellular process and antibiotic resistance in a bacte ferase is MurA . rium , comprising contacting the bacterium with an antisense In certain embodiments , the protein associated with a oligomer and /or a pharmaceutical composition described biochemical pathway and / or cellular process is a ribosomal herein . protein . In some embodiments , the ribosomal protein is a In certain embodiments , the bacterium is in a subject, and 50S ribosomal protein L28 . 30 the method comprises administering the antisense oligomer In certain embodiments, the 50S ribosomal protein L28 is to the subject. In some embodiments , the bacterium is RpmB . In particular embodiments , the protein associated selected from the genera Escherichia and Acinetobacter . In with a biochemical pathway and /or cellular process is a particular embodiments , the bacterium is an antibiotic -re cellular energy homeostasis protein . sistant strain of Escherichia or Acinetobacter . In some In certain embodiments , the cellular energy homeostasis 35 embodiments , the bacterium is a multi - drug resistant (MDR ) protein is an adenylate kinase . In specific embodiments , the strain of Escherichia or Acinetobacter . In specific embodi adenylate kinase is Adk . ments, the bacterium is Escherichia coli or Acinetobacter In certain embodiments , the protein associated with a baumannii. biochemical pathway and/ or cellular process is a protein In certain embodiments , the protein associated with a biosynthesis protein . In some embodiments , the protein 40 biochemical pathway and / or cellular process is an acyl biosynthesis protein is a translation initiation factor. In carrier protein . In some embodiments , the bacterium is various embodiments , the translation initiation factor is Escherichia coli and the acyl carrier protein is an AcpP InfA . protein encoded by acpP . In certain embodiments , the bac In certain embodiments , the protein associated with a terium is Acinetobacter baumannii and where the acyl biochemical pathway and/ or cellular process is a cell divi- 45 carrier protein is an Acpp protein encoded by acpP . sion protein . In particular embodiments , the cell division In some embodiments , the bacterium is Acinetobacter protein is a protein that assembles into a ring at the future spp . and the protein associated with a biochemical pathway site of the septum of bacterial cell division . and / or cellular process is a carboxyltransferase alpha subunit In some embodiments , the protein that assembles into a of an acetyl Coenzyme A carboxylase encoded by accA . ring at the future site of the septum of bacterial cell division 50 In certain embodiments , the bacterium is Escherichia coli is FtsZ . In certain embodiments , the protein associated with and the protein associated with a biochemical pathway a biochemical pathway and /or cellular process is an RNA and / or cellular process is a UDP - N - acetylglucosamine synthesis protein . 1 - carboxyvinyltransferase encoded by murA . In certain embodiments , the RNA synthesis protein is a In some embodiments, the bacterium is Escherichia coli sigma D factor of RNA polymerase. In particular embodi - 55 and the protein associated with a biochemical pathway ments , the sigma D factor of RNA polymerase is Rpod . and / or cellular process is a ribosomal protein encoded by In some embodiments , the protein associated with a rpmB . biochemical pathway and /or cellular process is an aromatic In particular embodiments , the bacterium is Escherichia compound biosynthesis protein . In some embodiments , the coli and the protein associated with a biochemical pathway aromatic compound biosynthesis protein is a chorismate 60 and /or cellular process is an adenylate kinase encoded by synthase ( 5 - enolpyruvylshikimate- 3 - phosphate phospho - adk . lyase ) . In particular embodiments , the chorismate synthase In some embodiments , the bacterium is Escherichia coli ( 5 - enolpyruvylshikimate - 3 - phosphate phospholyase ) is and the protein associated with a biochemical pathway AroC . and / or cellular process is a translation initiation factor In specific embodiments , the targeting sequence is set 65 encoded by infA . forth in SEQ ID NOS : 12 - 53 , comprises a fragment of at In certain embodiments, the bacterium is Acinetobacter least 10 contiguous nucleotides of SEQ ID NOS: 12 -53 , or spp . and the protein associated with a biochemical pathway US 10 ,391 , 098 B2 and / or cellular process is a protein that assembles into a ring In particular embodiments , the B -lactam antibiotic is at the future site of the septum of bacterial cell division selected from at least one of carbapenems, penicillin deriva encoded by ftsz . tives (penams ) , cephalosporins (cephems ) , and mono In some embodiments , the bacterium is Acinetobacter bactams. spp . and the protein associated with a biochemical pathway 5 In some embodiments , the bacterium is Acinetobacter and/ or cellular process is a sigma D factor of RNA poly spp . , the protein associated with a biochemical pathway merase encoded by rpoD . and /or cellular process is a carboxyltransferase alpha subunit In some embodiments , the bacterium is Acinetobacter of an acetyl Coenzyme A carboxylase encoded by accA , and spp . and the protein associated with a biochemical pathway the antimicrobial agent is selected from at least one of and / or cellular process is a chorismate synthase ( 5 -enolpyru - aminoglycoside antibiotics , tetracycline antibiotics , and vylshikimate - 3 - phosphate phospholyase ) encoded by aroC . B - lactam antibiotics . In particular embodiments , the bacterium is Escherichia In some embodiments , the bacterium is Escherichia coli , coli or Acinetobacter baumannii and the protein associated the protein associated with a biochemical pathway and /or with antibiotic resistance is selected from at least one of 15 cellular process is a UDP - N -acetylglucosamine 1 - car BlaT, Cml, and AdeA . boxyvinyltransferase encoded by murA , and the antimicro Somemethods comprise administering the oligomer sepa - bial agent is selected from at least one of aminoglycoside rately or concurrently with an antimicrobial agent, option - antibiotics, tetracycline antibiotics, and ß - lactam antibiotics . ally where administration of the oligomer increases suscep In particular embodiments , the bacterium is Escherichia tibility of the bacterium to the antimicrobial agent. 20 coli or Acinetobacter spp ., the protein associated with a In certain embodiments , the antimicrobial agent is biochemical pathway and / or cellular process is a ribosomal selected from one or more of a B - lactam antibiotic , an protein encoded by rpmB , and the antimicrobial agent is aminoglycoside antibiotic , and a polymyxin . selected from at least one of aminoglycoside antibiotics, In some embodiments , the B - lactam antibiotic is selected tetracycline antibiotics , and B - lactam antibiotics. from at least one of carbapenems, penicillin derivatives 25 In certain embodiments , the bacterium is Escherichia coli , (penams ), cephalosporins (cephems ) , and monobactams. the protein associated with a biochemical pathway and /or In particular embodiments , the carbapenem is selected cellular process is an adenylate kinase encoded by adk , and from one or more of meropenem , imipenem , ertapenem , the antimicrobial agent is selected from at least one of doripenem , panipenem , biapenem , razupenem , tebipenem , aminoglycoside antibiotics, tetracycline antibiotics , and tomonenem in specific embodiments the 30 ß - lactam antibiotics . lenapenem , and tomopenem . In specific embodiments , the 30 In some embodiments , the bacterium is Escherichia coli , carbapenem is meropenem . the protein associated with a biochemical pathway and / or In certain embodiments , the aminoglycoside antibiotic is cellular process is a translation initiation factor encoded by selected from one or more of tobramycin , gentamicin , infA , and the antimicrobial agent is selected from at least kanamycin a , amikacin , dibekacin , sisomicin , netilmicin1 ; , 35 one of aminoglycoside antibiotics , tetracycline antibiotics , neomycin B , neomycin C , neomycin E (paromomycin ) , and and B - lactam antibiotics . streptomycin . In specific embodiments , the aminoglycoside In some embodiments , the bacterium is Acinetobacter antibiotic is tobramycin . spp . , the protein associated with a biochemical pathway In certain embodiments , the polymyxin is selected from and /or cellular process is a protein that assembles into a ring one or more of colistin (polymyxin E ) , polysporin , neo - 40 at the future site of the septum of bacterial cell division sporin , or polymyxin B . In specific embodiments , the poly - encoded by fts Z , and the antimicrobial agent is selected from myxin is colistin . at least one of aminoglycoside antibiotics, tetracycline anti In certain embodiments , the bacterium is Escherichia coli biotics , and B - lactam antibiotics. or Acinetobacter spp . that expresses Blat , and the antimi- In certain embodiments, the bacterium is Acinetobacter crobial agent is a ß - lactam antibiotic . In some embodiments , 45 spp . , the protein associated with a biochemical pathway the ( B - lactam antibiotic is selected from at least one of and / or cellular process is a sigma D factor of RNA poly meropenem , imipenem , ertapenem , doripenem , panipenem , merase encoded by rpoD , and the antimicrobial agent is biapenem , razupenem , tebipenem , lenapenem , tomopenem , selected from at least one of aminoglycoside antibiotics , cephalosporins ( cephems) , penicillin , penicillin derivatives tetracycline antibiotics , and B - lactam antibiotics. ( penams) and ampicillin . 50 In particular embodiments , the bacterium is Acinetobacter In particular embodiments, the bacterium is Escherichia spp . , the protein associated with a biochemical pathway coli or Acinetobacter spp . that expresses cml, and the and /or cellular process is a chorismate synthase ( 5 - enolpyru antimicrobial agent is chloramphenicol. In certain embodi vylshikimate - 3 -phosphate phospholyase ) encoded by aroC , ments, the bacterium is Escherichia coli or Acinetobacter and the antimicrobial agent is selected from at least one of spp . that expresses adeA , and where the antimicrobial agent 55 aminoglycoside antibiotics, tetracycline antibiotics, and is selected from at least one of aminoglycoside antibiotics , B - lactam antibiotics . tetracycline antibiotics , and ( B - lactam antibiotics . In some embodiments, the oligomer reduces the minimum In certain embodiments , the aminoglycoside antibiotic is inhibitory concentration (MIC ) of the antimicrobial agent selected from at least one of tobramycin , gentamicin , against the bacterium by at least about 10 % relative to the kanamycin a , amikacin , dibekacin , sisomicin , netilmicin , 60 antimicrobial agent alone . neomycin B , neomycin C , neomycin E ( paromomycin ), and in some embodiments , the oligomer increases the suscep streptomycin . tibility of the bacterium to the antimicrobial agent by at least In some embodiments , the tetracycline antibiotic is about 10 % relative to the antimicrobial agent alone . selected from at least one of tetracycline , chlortetracycline , In certain embodiments, the combination of oligomer and oxytetracycline , demeclocycline , lymecycline , meclocy - 65 the antimicrobial agent synergistically increases the suscep cline , methacycline , minocycline , rolitetracycline , and tibility of the bacterium to the antibiotic relative to the doxycyline. oligomer and /or the microbial agent alone. In particular US 10 , 391 ,098 B2 10 embodiments , the antimicrobial agent is selected from colis - reduced bacterial by an additional ~ 1 - log relative to the tin , meropenem , and tobramycin . acpP - targeted PPMO alone . Tobramycin and control PPMOS In certain embodiments , the antimicrobial agent and the ( either alone or in combination ) did not have this significant antisense oligomer are administered separately. In various of an effect on bacterial growth . embodiments , the antimicrobial agent and the antisense 5 FIG . 6 shows that increasing amounts of acpP - targeted oligomer are administered sequentially . In some embodi- PPMO # 7 significantly decreased the minimum inhibitory ments , the antimicrobial agent and the antisense oligomer concentration (MIC ) of colistin against a multi - drug - resis are administered concurrently . tant strain of Acinetobacter baumannii (AYE ) in a concen Also included are methods of treating a multi - drug - tration - dependent manner. resistant (MDR ) Acinetobacter baumannii or Escherichia 10 FIG . 7 shows that increasing amounts of acpP - targeted coli bacterial infection in a subject, comprising administer - PPMO # 7 significantly decreased the minimum inhibitory ing to the subject an antibiotic selected from one or more of concentration (MIC ) of meropenem against a multi -drug tobramycin , meropenem , and colistin , in combination with resistant strain of Acinetobacter baumannii (AYE ) in a an antisense oligomer described herein . Certain antisense concentration -dependent manner. oligomers comprise a targeting sequence of sufficient length 15 FIG . 8 shows that increasing amounts of acpP - targeted and complementarity to specifically hybridize to an mRNA PPMO # 7 significantly decreased the minimum inhibitory target sequence of a bacterial acpP gene that encodes an acyl concentration (MIC ) of tobramycin against a multi - drug carrier protein (AcpP ) , where the combination of the anti - resistant strain of Acinetobacter baumannii (AYE ) in a sense oligomer and the antibiotic synergistically increases concentration - dependent manner. the susceptibility of the MDR Acinetobacter or the MDR 20 FIGS . 9A - 9C show MIC comparisons of E . coli and Escherichia coli to the antibiotic relative to the antibiotic Acinetobacter strains in rich and minimal media with vari alone . ous PPMOs and Scr controls . FIG . 9A shows the results for E . coli W3110 challenged with acpP (PPMO # 1) , acpP BRIEF DESCRIPTION OF THE FIGURES (PPMO # 2 ), acpP (PPMO # 3 ), acpP (PPMO # 4 ), acpP 25 (PPMO # 5 ), murA (PPMO # 24 ), and Scramble (Scr ) controls . FIG . 1A shows an exemplary morpholino oligomer struc - FIGS. 9B - 9C shows the results for A . baumannii AYE (9B ) ture with a phosphorodiamidate linkage . FIGS . 1B - E show and A . baumannii 0057 (9C ) challenged with acpP the repeating subunit segment of exemplary morpholino (PPMO # 7 ) , acpP (PPMO # 8 ) , acpP ( PPMO # 13 ) , acpP oligomers , designated B through E . FIGS. 1F - H show exem (PPMO # 14 ) , ftsZ (PPMO # 33 ) , ftsZ ( PPMO # 34 ), rpmB plary peptide PMO conjugates structures used in the exem - 30 (PPMO # 29 ) , and Scramble ( Scr) controls . The bacteria were plary PPMOs . challenged in either MHII (black bar ) or AB MinimalMedia FIG . 2A shows that acpP - targeted PPMO # 2 not only ( grey bar) . reduced bacterial growth ( colony - forming units ; CFUs ) of a FIGS. 10A - 10C show the kinetics of minimal bactericidal multi - drug - resistant strain of E . coli by about 1 - log relative (MBC ) viability assays on the growth of E . coli 1101851, A . to scramble PPMO control , but in combination with 35 baumannii AYE and 0057 upon challenge with PPMOs tobramycin also synergistically reduced bacterial growth by targeted against acpP . The cultures were grown aerobically over 4 - logs relative to controls . Tobramycin and control at 37° C . with various concentrations of PPMO , as indicated . PPMOs ( either alone or in combination ) had no significant Samples were taken at the times noted , back -diluted 150 effect on bacterial growth . FIG . 2B ( linear scale ) and FIG . mM NaCl, and plated on blood agar. The plates were 2C ( log scale ) show that increasing amounts of acpP - 40 incubated for 18 hours and the resulting colony counts were targeted PPMO # 1 significantly decreased the minimum used to determine the CFU /mL . FIG . 10A shows the results inhibitory concentration (MIC ) of tobramycin against a for E . coli 1101851 challenged with PPMOs acpP (PPMO # 3 ) multi - drug - resistant strain of E . coli in a concentration and ( RFR ) 4 -Scramble (Scr ) control. FIGS . 10B - 10C respec dependent manner. tively show the results for A . baumannii AYE and AB0057 FIG . 3 shows that acpP - targeted PPMO # 7 not only 45 challenged with PPMOs acpP (PPMO # 7 ) and (RXR ) 4 reduced bacterial growth ( colony - forming units ; CFUS) of a Scramble (Scr ) control. multi - drug - resistant strain of Acinetobacter baumannii FIGS . 11A - 11L show Transmission Electron Microscopy (AYE ) by about 6 -logs relative to scramble PPMO control, ( TEM ) of thin -sectioned A . baumannii AYE . FIGS. 11A but in combination with colistin also synergistically reduced 11B show A . baumannii AYE before treatment with PPMOs bacterial growth to undetectable levels (an additional ~ 3 50 and FIGS. 116 - 11H shows after a 6 hour incubation alone . logs relative to the acpP - targeted PPMO alone ) . Colistin and FIGS . 11C - 11D show the results for 40 UM scramble control control PPMOs ( either alone or in combination ) did not have at 0 hour and FIGS . 111- 11J show the results for 40 uM this significant of an effect on bacterial growth . scramble control at 6 hours treatment. FIGS . 11E - 11F show FIG . 4 shows that acpP - targeted PPMO # 7 not only the results for 40 uM acpP at 0 hour and FIGS . 11K - 11L reduced bacterial growth (colony - forming units ; CFUs) of a 55 show the results for 40 uM at 6 hours treatment. CW , cell multidrug - resistant strain of Acinetobacter baumannii wall section ; arrows point out cell wall disruption . ( AYE ) by about 5 - logs relative to scramble PPMO control, FIGS . 12A - 12F show the synergy between acpP -targeted but in combination with meropenem also synergistically PPMOs and three different antibiotics against the multidrug reduced bacterial growth by an additional ~ 2 -logs relative to resistant E . coli strain AIS070834 . The MIC of colistin , the acpP - targeted PPMO alone . Meropenem and control 60 meropenem , and tobramycin was measured with various PPMOs (either alone or in combination ) did not have this concentrations of acpP - targeted PPMO (PPMO # 1) or significant of an effect on bacterial growth . scrambled (Scr ) control PPMO . Viable cells were counted in FIG . 5 shows that acpP -targeted PPMO # 7 not only 24 - hour cultures with antibiotic alone, PPMO alone, or in reduced bacterial growth ( colony - forming units ; CFUs ) of a combination thereof . FIGS. 12A - 12B show the results for multi - drug- resistant strain of Acinetobacter baumannii 65 colistin , FIGS. 12C - 12D show the results for meropenem , (AYE ) by about 6 - logs relative to scramble PPMO control, and FIGS . 12E - 12F show the results for tobramycin . In all but in combination with tobramycin also synergistically instances, the acpP - targeted PPMO significantly reduced the US 10 , 391 , 098 B2 12 MIC of the tested antibiotics . The free peptide (RXR ) 4XB step or element or group of steps or elements but not the was also tested and by itself had an unmeasurable MIC (data exclusion of any other step or element or group of steps or not shown ) . Error bars indicate standard deviation ( N = 2 for elements . all experiments ) . By " consisting of” is meant including , and limited to , FIGS . 13A - 13D show the MICs of PPMOs targeted 5 whatever follows the phrase " consisting of: ” Thus , the against various genes in selected bacterial strains . FIG . 13A phrase " consisting of” indicates that the listed elements are shows the results for E . coli strains grown in MHII. FIG . required or mandatory , and that no other elements may be 13B shows the results for E . coli strains grown in MOPS present. By “ consisting essentially of” is meant including minimal media . FIG . 13C shows the results for Acineto - any elements listed after the phrase , and limited to other bacter species grown in MHII. FIG . 13D shows the results 10 elements that do not interfere with or contribute to the for Acinetobacter species grown in AB minimal media . activity or action specified in the disclosure for the listed FIGS . 14A - 14F show synergy of ftsZ PPMO with 3 elements . Thus, the phrase " consisting essentially of” indi antibiotics . The MIC of classic antibiotics (FIG . 14A , FIG . cates that the listed elements are required or mandatory , but 14C , FIG . 14E ) was measured with various concentrations that other elements are optional and may or may not be of ftsZ PPMO (PPMO # 46 ) or scrambled ( Scr ) PPMO (Scr - 15 present depending upon whether or not they materially affect 1 ) , using as indicator the multidrug resistant E . coli the activity or action of the listed elements . AIS070834 . Viable cells were counted in 24 - h cultures with As used herein , the terms " contacting a cell” , “ introduc antibiotic or PPMO alone, and in combinations where syn - ing ” or “ delivering ” include delivery of the oligomers ergy was apparent ( FIG . 14B , FIG . 14D , FIG . 14F ) . The free described herein into a cell by methods routine in the art, peptide (RXR ) XB was also tested for synergy with the 20 e . g . , transfection ( e . g . , liposome, calcium - phosphate , poly antibiotics , although by itself had an unmeasurable MIC ethyleneimine ), electroporation (e .g ., nucleofection ), micro ( data not shown ). Error bars indicate standard deviation . injection ), transformation , and administration . N = 2 for all experiments . The terms " cell penetrating peptide ” (CPP ) or “ a peptide FIGS. 15A - 15B show the minimum inhibitory concentra - moiety which enhances cellular uptake ” are used inter tion (MIC ) of classical antibiotics with added PPMOs that 25 changeably and refer to cationic cell penetrating peptides , target resistance genes in E . coli SMS- 3 - 5 . ( FIG . 15A ) MIC also called “ transport peptides” , “ carrier peptides ” , or “ pep of ampicillin with blaT - (RXR ) XB ( PPMO # 66 ) . ( FIG . 15B ) tide transduction domains ” . In some aspects, the peptides MIC of chloramphenicol with cm1A - (RXR ) XB have the capability of inducing cell penetration within about (PPMO # 67 ) . N = 3 . or at least about 30 % , 40 % , 50 % , 60 % , 70 % , 80 % , 90 % , or FIGS. 16A - 16B show that PPMO mediated knockdown 30 100 % of cells of a given population and / or allow macro of nonessential resistance genes shows recovery of classical molecular translocation to or within multiple tissues in vivo antibiotics efficacy. The graphs represent MICs performed upon systemic administration . Particular examples of CPPs by diluting PPMOs (ade A -( RXR ) XB (PPMO # 65 ) ; Scr include “ arginine- rich peptides. ” CPPs are well -known in (RXR ) XB (Scr - 2 ) ; and Peptide (RXR ) . XB , not shown ) the art and are disclosed , for example , in U . S . Application independently and with tobramycin in A . baumannii strain 35 No . 2010 / 0016215 and International Patent Application AYE in both ( FIG . 16A ) nutrient rich and ( FIG . 16B ) Publication Nos . WO 2004 /097017 , WO 2009 /005793 , and minimal medias. N = 3 . WO 2012 / 150960 , all of which are incorporated by refer ence in their entirety . DETAILED DESCRIPTION “ An electron pair ” refers to a valence pair of electrons that 40 are not bonded or shared with other atoms. I . Definitions “ Homology ” refers to the percentage number of amino Unless defined otherwise , all technical and scientific acids that are identical or constitute conservative substitu terms used herein have the same meaning as commonly tions . Homology may be determined using sequence com understood by those of ordinary skill in the art . Although any parison programs such as GAP (Deveraux et al. , 1984 , methods and materials similar or equivalent to those 45 Nucleic Acids Research 12 , 387 - 395 ) or BLAST. In this way described herein can be used in the practice or testing of the sequences of a similar or substantially different length to present disclosure , preferred methods and materials are those cited herein could be compared by insertion of gaps described . For the purposes of the present disclosure , the into the alignment , such gaps being determined , for following terms are defined below . example , by the comparison algorithm used by GAP . The articles " a " and " an " are used herein to refer to one 50 By “ isolated ” is meant material that is substantially or or to more than one ( i .e . , to at least one of the grammatical essentially free from components that normally accompany object of the article . By way of example , " an element ” it in its native state . For example , an “ isolated polynucle means one element or more than one element. otide ” or “ isolated oligomer, ” as used herein ,may refer to a By “ about ” is meant a quantity , level , value, number, polynucleotide that has been purified or removed from the frequency , percentage , dimension , size , amount, weight, or 55 sequences that flank it in a naturally - occurring state , e . g . , a length that varies by as much as 30 , 25 , 20 , 15 , 10 , 9 , 8 , 7 , DNA fragment that is removed from the sequences that are 6 , 5 , 4 , 3 , 2 or 1 % to a reference quantity , level , value , adjacent to the fragment in the genome. The term “ isolating ” number, frequency, percentage , dimension , size , amount, as it relates to cells refers to the purification of cells ( e . g ., weight, or length . fibroblasts , lymphoblasts ) from a source subject ( e . g . , a By " coding sequence” is meant any nucleic acid sequence 60 subject with a polynucleotide repeat disease ) . In the context that contributes to the code for the polypeptide product of a of mRNA or protein , “ isolating ” refers to the recovery of gene . By contrast , the term “ non - coding sequence ” refers to mRNA or protein from a source , e . g ., cells . any nucleic acid sequence that does not directly contribute The term “ modulate ” includes to “ increase” or “ decrease ” to the code for the polypeptide product of a gene. one or more quantifiable parameters , optionally by a defined Throughout this specification , unless the context requires 65 and/ or statistically significant amount. By " increase ” or otherwise , the words “ comprise , " " comprises , ” and “ com - “ increasing, " " enhance " or " enhancing , " or " stimulate ” or prising ” will be understood to imply the inclusion of a stated " stimulating, ” refers generally to the ability of one or US 10 , 391, 098 B2 13 14 antisense compounds or compositions to produce or cause a As used herein , “ nucleobase ” (Nu ), “ base pairing moiety ” greater physiological response ( i. e ., downstream effects ) in or “ base” are used interchangeably to refer to a purine or a cell or a subject relative to the response caused by either pyrimidine base found in native DNA or RNA (uracil , no antisense compound or a control compound . Relevant thymine , adenine, cytosine, and guanine ) , as well as analogs physiological or cellular responses ( in vivo or in vitro ) will 5 of the naturally occurring purines and pyrimidines , that be apparent to persons skilled in the art . An “ increased ” or confer improved properties , suchsuch as binding affinity to the “ enhanced ” amount is typically a “ statistically significant” oligomer. Exemplary analogs include hypoxanthine ( the amount, and may include an increase that is 1. 1 , 1 .2 , 2 , 3, 4 , base component of the nucleoside inosine ); 2 ,6 -diaminopu 5 , 6 , 7 , 8 , 9 , 10 , 15 , 20 , 30 , 40 , 50 or more times (e . g. , 500 , 1000 times ) ( including all integers and ranges between and 10 rine0 ; 5 -methyl cytosine ; C5 -propynyl - modified pyrimidines; above 1 ) , e . g . , 1 . 5 , 1 . 6 , 1 . 7 . 1 . 8 ) the amount produced by no 9 - ( aminoethoxy ) phenoxazine ( G - clamp) and the like . antisense compound ( the absence of an agent ) or a control A nucleobase covalently linked to a ribose , sugar analog compound . The term " reduce ” or “ inhibit ” may relate gen or morpholino comprises a nucleoside . “ Nucleotides” are erally to the ability of one or more antisense compounds or composed of a nucleoside together with one phosphate compositions to “ decrease” a relevant physiological or cel - 15 group . The phosphate groups covalently link adjacent lular response , such as a symptom of a disease or condition nucleotides to one another to form an oligomer. described herein . as measured according to routine tech - An oligomer “ specifically hybridizes ” to a target sequence niques in the diagnostic art . Relevant physiological or cel- if the oligomer hybridizes to the target under physiological lular responses ( in vivo or in vitro ) will be apparent to conditions , with a Tm substantially greater than 40° C . or persons skilled in the art , and may include reductions in 20 45° C . , preferably at least 50° C . , and typically 60° C . - 80° bacterial cell growth , reductions in the minimum inhibitory C . or higher. Such hybridization preferably corresponds to concentration (MIC ) of an antimicrobial agent, and others . A stringent hybridization conditions . At a given ionic strength “ decrease” in a response may be “ statistically significant” as and pH , the Tm is the temperature at which 50 % of a target compared to the response produced by no antisense com sequence hybridizes to a complementary polynucleotide . pound or a control composition , and may include a 1 % , 2 % , 25 Such hybridization may occur with “ near ” or “ substantial” 3 % , 4 % , 5 % , 6 % , 7 % , 8 % , 9 % , 10 % , 11 % , 12 % , 13 % , 14 % , complementarity of the antisense oligomer to the target 15 % , 16 % , 17 % , 18 % , 19 % , 20 % , 25 % , 30 % , 35 % , 40 % , sequence , as well as with exact complementarity . 45 % , 50 % , 55 % , 60 % , 65 % , 70 % , 75 % , 80 % , 85 % , 90 % , As used herein , “ sufficient length ” includes an antisense 95 % , or 100 % decrease , including all integers and ranges in oligomer that is complementary to at least about 8 , more between . 30 typically about 8 -10 , 8 - 11, 8 - 12 , 8 - 13 , 8 - 14 , 8 -15 , 8 - 16 , As used herein , an " antisense oligomer ," " oligomer, " or 8 - 17 , 8 - 18 , 8 - 19 , 8 - 20 , 8 - 30 , 8 -40 , or 10 - 11 , 10 - 12 , 10 - 13 , " oligomer” refers to a linear sequence of nucleotides, or 10 - 14 , 10 - 15, 10 - 16 , 10 -17 , 10 -18 , 10 - 19 , 10 - 20 , 10 - 30 , nucleotide analogs, which allows the nucleobase to hybrid - 10 - 40 ( including all integers and ranges in between ) con ize to a target sequence in an RNA by Watson - Crick base tiguous or non -contiguous nucleobases in a region of a pairing , to form an oligomer :RNA heteroduplex within the 35 bacterial mRNA target sequence . An antisense oligomer of target sequence. The terms" antisense oligomer ," " antisense sufficient length has at least a minimal number of nucleo oligomer, " " oligomer, ” and “ compound” may be used inter - tides to be capable of specifically hybridizing to a region of changeably to refer to an oligomer. The cyclic subunits may the bacterial mRNA target . Preferably an oligomer of suf be based on ribose or another pentose sugar or , in certain ficient length is from 8 to 30 nucleotides in length , for embodiments , a morpholino group ( see description of mor - 40 example , about 10 - 20 nucleotides in length . pholino oligomers below ) . The terms “ sequence identity ” or, for example , compris The term “ oligomer, " " oligomer, " or " antisense oligomer ” ing a “ sequence 50 % identical to , " as used herein , refer to also encompasses an oligomer having one or more addi- the extent that sequences are identical on a nucleotide -by tional moieties conjugated to the oligomer , e . g . , at its 3 ' - or nucleotide basis or an amino acid -by - amino acid basis over 5 ' - end , such as a polyethylene glycolmoiety or other hydro - 45 a window of comparison . Thus , a " percentage of sequence philic polymer, e . g . , one having 10 - 100 monomeric sub - identity ” may be calculated by comparing two optimally units , which may be useful in enhancing solubility, or a aligned sequences over the window of comparison , deter moiety such as a lipid or peptide moiety that is effective to mining the number of positions at which the identical enhance the uptake of the compound into target bacterial nucleic acid base ( e . g . , A , T , C , G , I ) or the identical amino cells and /or enhance the activity of the compound within the 50 acid residue (e .g . , Ala , Pro , Ser , Thr, Gly , Val , Leu , Ile , Phe , cell, e . g . , enhance its binding to a target polynucleotide . Tyr, Trp , Lys , Arg , His , Asp , Glu , Asn , Gin , Cys and Met ) A “ nuclease -resistant ” oligomers refers to one whose occurs in both sequences to yield the number of matched backbone is substantially resistant to nuclease cleavage , in positions , dividing the number of matched positions by the non - hybridized or hybridized form ; by common extracellu total number of positions in the window of comparison ( i . e . , lar and intracellular nucleases in the body or in a bacterial 55 the window size ) , and multiplying the result by 100 to yield cell ( for example , by exonucleases such as 3 ' - exonucleases , the percentage of sequence identity . Optimal alignment of endonucleases , RNase H ) ; that is , the oligomer shows little sequences for aligning a comparison window may be con or no nuclease cleavage under normal nuclease conditions to ducted by computerized implementations of algorithms which the oligomer is exposed . A “ nuclease -resistant het - (GAP , BESTFIT, FASTA , and TFASTA in the Wisconsin eroduplex ” refers to a heteroduplex formed by the binding of 60 Genetics Software Package Release 7 . 0 , Genetics Computer an antisense oligomer to its complementary target , such that Group , 575 Science Drive Madison , Wis ., USA ) or by the heteroduplex is substantially resistant to in vivo degra - inspection and the best alignment (i .e . , resulting in the dation by intracellular and extracellular nucleases, which are highest percentage homology over the comparison window ) capable of cutting double - stranded RNA/ RNA or RNA generated by any of the various methods selected . Reference DNA complexes . A " heteroduplex ” refers to a duplex 65 also may be made to the BLAST family of programs as for between an antisense oligomer and the complementary example disclosed by Altschul et al. , Nucl. Acids Res . portion of a target RNA . 25 : 3389 , 1997 . US 10 , 391 , 098 B2 15 16 A “ subject ” or a “ subject in need thereof includes a A “ targeting sequence” may have " near” or “ substantial ” mammalian subject such as a human subject. complementarity to the target sequence and still function for The terms “ TEG , ” “ EG3, ” or “ triethylene glycol tail” the purpose of the present disclosure , that is , still be refer to triethylene glycol moieties conjugated to the oli - “ complementary. ” Preferably , the oligomer analog com gomer , e . g . , at its 3 ' - or 5 - end . For example , in some 5 pounds employed in the present disclosure have at most one embodiments , “ TEG ” includes , for example , wherein T of mismatch with the target sequence out of 10 nucleotides, and preferably at most onemismatch outof 20 . Alternatively , the the compound of formula ( I ) , ( II ) , or (III ) is of the formula : antisense oligomers employed have at least 90 % sequence homology, and preferably at least 95 % sequence homology , 10 with the exemplary targeting sequences as designated HO herein . Hotte As used herein , the term " quantifying ” , “ quantification ” or other related words refer to determining the quantity , mass, or concentration in a unit volume, of a nucleic acid , polynucleotide , oligomer, peptide, polypeptide , or protein . As used herein , " treatment" of a subject ( e . g . a mammal , such as a human ) or a cell is any type of intervention used O = P in an attempt to alter the natural course of the individual or cell . Treatment includes , but is not limited to , administration 20 of a pharmaceutical composition , and may be performed either prophylactically or subsequent to the initiation of a pathologic event or contact with an etiologic agent . Also included are " prophylactic ” treatments , which can be The term “ pip -PDA ” refers to a 5 ' terminal piperazine directed to reducing the rate of progression of the disease or phosphorodiamidate moiety that connects a G group , where 25 condition being treated , delaying the onset of that disease or condition , or reducing the severity of its onset. “ Treatment” the G group comprises a cell -penetrating peptide (CPP ) and or “ prophylaxis ” does not necessarily indicate complete linker moiety further discussed below , to the 5 ' end of the eradication , cure , or prevention of the disease or condition , oligomer by way of an amide bond between the G group or associated symptoms thereof. linker and the piperazinyl nitrogen . For example , in some 30 II. Bacterial Targeting Sequences embodiments , “ pip -PDA ” includes wherein T of the com Certain embodiments relate to antisense oligomers, and pound of formula ( I ) or ( II) is of the formula : related compositions and methods, which are of sufficient length and complementarity to specifically hybridize to a

9 bacterial mRNA target sequence that encodes a gene in a

- 35 biochemical pathway and / or cellular process . General Z examples include: cell division , global gene regulatory mechanisms, fatty acid biosynthesis , ribosomal proteins, DNA replication , transcription , translation initiation , lipopo lysaccharide biosynthesis , peptidoglycan biosynthesis , 40 nucleic acid biosynthesis , intermediary metabolism , and O = P - N (CH3 ) 2 antibiotic resistance . Particular examples of genes in bio chemical pathways and cellular processes include: rpsi and rpmB ( ribosomal proteins ) ; IpxC , waac , waag , waaA , waaF , IpxA , and IpxB ( lipopolysaccharide biosynthesis ) ; min 45 murA ( formerly known as murZ ) , mra Y , murC , murB , murE , murF , and murG (peptidoglycan biosynthesis ) ; fabG , acpp , The term “ target sequence” refers to a portion of the target accA , accB , and fabZ ( fatty acid biosynthesis) ; adk (cellular RNA , for example, a bacterial mRNA, against which the energy homeostasis ) ; infÀ ( transcription antitermination antisense oligomer is directed , that is , the sequence to which and /or protein synthesis ) ; ftsZ ( cell division ) ; rpoD (RNA the oligomer will hybridize by Watson - Crick base pairing of 50 synthesis ) ; aroC ( aromatic compound biosynthesis ) . a complementary sequence . In certain embodiments , the Examples of antibiotic resistance genes include blat , cml, target sequence may be a contiguous region of the transla - and adeA . In some embodiments , themRNA target sequence tion initiation region of a bacterial gene . that encodes the gene is from Acinetobacter , e . g . , Acineto The term “ targeting sequence ” or “ antisense targeting bacter baumannii . In some embodiments , the mRNA target sequence” refers to the sequence in an oligomer that is 55 sequence that encodes the gene is from Escherichia , e . g . , E . complementary or substantially complementary to the target coli . sequence in the RNA , e. g ., the bacterial mRNA . The entire In some embodiments , the bacterial target is a gene or sequence , or only a portion , of the antisense compound may protein that is associated with biosynthesis of fatty acids . be complementary to the target sequence . For example , in an General examples of proteins associated with fatty acid oligomer of about 10 - 30 bases , about 6 , 7 , 8 , 9 , 10 , 11 , 12 , 60 biosynthesis include : acyl carrier protein ( ACP ), such as 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21, 22 , 23 , 24 , 25 , 26 , 27 , 28 , AcpP , that plays an essential role in stabilizing and shuttling or 29 of the bases may be targeting sequences that are the intermediate fatty acid chain to each of the enzymes in complementary to the target region . Typically , the targeting the fatty acid synthase complex ; acyl carrier protein syn sequence is formed of contiguous bases , but may alterna - thase (AcpS ) , an enzyme that transfers the 4 '- phosphopan tively be formed of non -contiguous sequences that when 65 tetheine prosthetic group to apo -ACP to form the functional placed together , e .g ., from opposite ends of the oligomer, holo - ACP ; acetyl -CoA carboxylase , an enzyme composed constitute sequence that spans the target sequence . of four proteins that catalyzes the conversion of acetyl- CoA US 10 ,391 , 098 B2 17 18 to malonyl- CoA in the first committed step of fatty acid In some embodiments , the bacterial target is a gene or biosynthesis : AccA (carboxyltransferase alpha subunit cata protein that is associated with cell division . A particular lyzing the transfer of the carboxyl group from biotin to example of a gene associated with cell division includes a acetyl - CoA to form malonyl - CoA ), AccB (biotin carboxyl ftsZ gene , which encodes a protein that assembles into a ring carrier protein , BCCP, carrying the biotin prosthetic group 5 at the future site of the septum of bacterial cell division . This covalently attached to a lysine residue proximal to the is a prokaryotic homologue to the eukaryotic protein tubulin . carboxyl terminus) , AccC (biotin carboxylase catalyzing the In some embodiments , the bacterial target is a gene or protein that is associated with RNA synthesis . A particular carboxylation of protein bound biotin with bicarbonate ), example of a gene associated with RNA synthesis includes AccD ( carboxyltransferase beta subunit catalyzing the trans 10 an rpoD gene, which encodes a sigma D ( sigma 70 ) factor fer of the carboxyl group from biotin to acetyl- CoA to form of RNA polymerase that allows binding of the polymerase to malonyl- CoA ); fatty acid biosynthesis ( Fab ) enzymes, such gene promoters and is important for transcribing most genes as FabA , Fabl, FabF , FabB , FabD , FabH , FabG and FabZ , in growing cells . Genes recognized by this sigma factor have that each catalyze either elongation or tailoring steps on the promoter consensus sequences centered at 10 and 35 nucleo growing fatty acid chain . Particular examples of genes 15 fides before the start of transcription . associated with fatty acid biosynthesis include acpP and the The biosynthesis of aromatic compounds is important for carboxyltransferase alpha subunit accA . An exemplary the growth and survival of bacterial cells . The shikimate translational start codon region sequence of the acyl carrier pathway is a biosynthetic route in microorganisms that lead protein acpP gene is provided in Table 1 below . to the synthesis of chorismic acid , a central precursor for Specific embodiment therefore relate to antisense oligom - 20 other aromatic compounds . In some embodiments, the bac ers , and related compositions and methods, which are of terial target is a gene or protein that is associated with sufficient length and complementarity to specifically hybrid - aromatic compound biosynthesis . A particular example of a ize to an mRNA target sequence of a bacterial acpP gene, gene associated with aromatic compound biosynthesis which encodes an acyl carrier protein (ACP ). In some includes an aroC gene , which encodes chorismate synthase embodiments , the acpP gene is from Acinetobacter , e . g ., 25 ( 5 - enolpyruvylshikimate - 3 - phosphate phospholyase ) , the Acinetobacter baumannii . In some embodiments , the acpp final enzyme in the shikimate pathway that catalyzes the gene is from Escherichia , e . g ., E . coli. conversion of 5 - enolpyruvylshikimate - 3 - phosphate to cho The bacterial cell wall peptidoglycan is an essential rismic acid . cellular component involved in the maintenance of shape In some embodiments , the gene or protein is associated and protection from osmotic shock lysis . The Escherichia 30 with resistance of the bacteria to at least one antimicrobial coli peptidoglycan is assembled from a basic building block agent, i . e . , an antibiotic resistance gene . General examples composed of N -acetylglucosamine (GlcNAc ) and N -acetyl - of antibiotic resistance genes include beta - lactamases, which muramic acid with an attached pentapeptide . In some can enzymatically deactivate certain antimicrobial agents , embodiments , the bacterial target is a gene or protein that is and proteins that increase the permeability or active efflux associated with peptidoglycan biosynthesis . A particular 35 (pumping - out ) of an antimicrobial agent. Particular example of a gene associated with peptidoglycan biosyn - examples of antibiotic resistance genes include TEM beta thesis include mura ( formerly known as murZ ) , which lactamase (blaT ) , chloramphenicol resistance gene (cml ) , encodes a UDP - N -acetylglucosamine 1 -carboxyvinyltrans - and resistance -nodulation - cell division ( RND ) - type multi ferase , which catalyzes the first committed step of peptido - drug efflux pump subunit AdeA ( adeA ) . glycan biosynthesis . The enzyme catalyzes the transfer of 40 In certain embodiments , the target sequence contains all enolpyruvate from phosphoenolpyruvate to the 3 -OH of or a portion ( e . g . , 1 or 2 nucleotides ) of a translational start UDP - N -acetylglucosamine . codon of the bacterial mRNA . In some embodiments , the The ribosome is crucial for translation of mRNA mol target sequence contains a sequence that is about or within ecules into proteins . In some embodiments , the bacterial about 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , target is a gene or protein that is associated with ribosomal 45 18, 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 bases proteins . A particular example of a gene associated with upstream or downstream of a translational start codon of the ribosomal proteins is rpmB , a 50S ribosomal protein L28 bacterial mRNA target sequence , including common and essential for ribosome assembly and translation . alternative start codons ( e . g ., AUG , GUG , UUG , AUU , In some embodiments , the bacterial target is a gene or CUG ) . For example , in particular embodiments , the 5 ' -end protein that is associated with cellular energy homeostasis . 50 of the target sequence is the adenine, uracil , or guanine A particular example of a gene associated with cellular nucleotide ( respectively ) in an AUG start codon of the energy homeostasis includes an adenylate kinase ( adk ) gene, bacterial mRNA . In some embodiments, the 5 '- end of the which encodes a phosphotransferase enzyme that catalyzes target sequence is the guanine , uracil, or guanine nucleotide the interconversion of adenine nucleotides . (respectively ) in a GUG start codon of the bacterial mRNA . In some embodiments , the bacterial target is a gene or 55 In some embodiments , the 5 ' - end of the target sequence is protein that is associated with transcription antitermination the uracil, uracil, or guanine nucleotide ( respectively ) in a and/ or protein biosynthesis. A particular example of a gene UUG start codon of the bacterial mRNA . In some embodi associated with transcription antitermination and /or protein ments , the 5 '- end or 3 - end of the target sequence begins at biosynthesis includes translation initiation factor IF1. IF1 , residue 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , encoded by infA , is a protein containing an S1 - like domain 60 18 , 19 , 20 , 21, 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , or 30 that may play a role in binding and melting nucleic acid downstream of the last ( third ) nucleotide of a translational secondary structure and transcription antitermination . Other start codon of the bacterial mRNA . In some embodiments , functions may also include increasing the rate of 70S the 5 '- end or 3 - end of the target sequence begins at residue ribosome dissociation and subunit association and involve - 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , ment in the fidelity of translation initiation through stimu - 65 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29, or 30 upstream of the lation of other translation initiation factor activities , such as first nucleotide of a translational start codon of the bacterial IF2 and IF3 . mRNA US 10 ,391 , 098 B2 19 20 The target sequences of exemplary target genes from 82 % , 83 % , 84 % , 85 % , 86 % , 87 % , 88 % , 89 % , 90 % , 91 % , Acinetobacter baumannii and E . coli are provided in Table 92 % , 93 % , 94 % , 95 % , 96 % , 97 % , 98 % , 99 % or 100 % 1 below . sequence complementarity , between the oligomer and the target sequence . Oligomer backbones that are less suscep TABLE 1 5 tible to cleavage by nucleases are discussed herein . Mis Exemplary Target Sequences matches, if present, are typically less destabilizing toward Table 1A : Exemplary Antibiotic the end regions of the hybrid duplex than in the middle . The Resistance Target Sequences number of mismatches allowed will depend on the length of the oligomer, the percentage of G : C base pairs in the duplex , SEO ID Description Sequence * and the position of the mismatch (es ) in the duplex , accord NO : ing to well understood principles of duplex stability . E . coli acpP Acyl AAAGCGAGTTTTGATAGG 69 carrier protein AAATTTAAGAGTATGAGC Although such an antisense oligomer is not necessarily (ACP ) ACTATCGAAGAACGCGTT 100 % complementary to the target sequence , it is effective AAGAAA 15 to stably and specifically bind to the target sequence , for A . baumannii TTTTAAAAATTTTTATAT 70 example , such that translation of the target RNA is reduced . acpP Acyl carrier TCAATTAAACTAGTGGCA The stability of the duplex formed between an oligomer protein ( ACP ) AATCAAACGCCACAAGCA and a target sequence is a function of the binding Tm and the ATGAGGAGAATTCCTGTG susceptibility of the duplex to cellular enzymatic cleavage . AGCGATATCGAACAACGC The Tm of an oligomer with respect to complementary * The thymines ( T ) can be uracils ( U ) , and vice versa sequence RNA may be measured by conventional methods , such as those described by Hames et al. , Nucleic Acid Thus, in certain embodiments , antisense targeting Hybridization , IRL Press, 1985 , pp . 107 - 108 or as described sequences are designed to hybridize to a region of one or in Miyada C . G . and Wallace R . B . , 1987 , Oligomer Hybrid more of the target sequences listed in Table 1 ( e .g . , SEQ ID NO : 69 or 70 ) or a target gene described herein . Selected 25 ization Techniques, Methods Enzymol. Vol. 154 pp . 94 - 107 . antisense targeting sequences can be made shorter, e .g ., In certain embodiments , antisense oligomers may have a about 8 , 9 , 10 , 11, 12 , 13 , 14 , or 15 bases, or longer, e . g . , binding Tm , with respect to a complementary - sequence about 20 , 30 , or 40 bases , and include a small number of RNA , of greater than body temperature and preferably mismatches , as long as the sequence is sufficiently comple greater than about 45° C . or 50° C . Tm ' s in the range 60 - 80° mentary to reduce transcription or translation upon hybrid - 30 C . or greater are also included . According to well -known ization to the target sequence , and optionally forms with the principles, the Tm of an oligomer , with respect to a comple RNA a heteroduplex having a Tm of 45° C . or greater. mentary -based RNA hybrid , can be increased by increasing In certain embodiments , the degree of complementarity the ratio of C : G paired bases in the duplex , and /or by between the target sequence and antisense targeting increasing the length ( in base pairs ) of the heteroduplex . At sequence is sufficient to form a stable duplex . The region of 35 the same time, for purposes of optimizing cellular uptake , it complementarity of the antisense oligomers with the target may be advantageous to limit the size of the oligomer. RNA sequence may be as short as 8 - 9 bases , 8 - 10 bases , Tables 2A - B below show exemplary targeting sequences 8 - 11 bases , 8 - 12 bases , 10 - 11 bases, 10 - 12 bases, but can be (in a 5 -to - 3 ' orientation ) of the antisense oligomers 12 - 15 bases or more , e . g ., 10 -40 bases, 12 - 30 bases , 12 - 25 described herei bases , 15 - 25 bases, 12 - 20 bases , or 15 - 20 bases , including12 :23 described herein . all integers in between these ranges. An antisense oligomer 40 of about 10 - 15 bases is generally long enough to have a TABLE 2A unique complementary sequence . In certain embodiments , a Exemplary Fatty Acid minimum length of complementary bases may be required to Biosynthesis - Associated Targeting achieve the requisite binding Tm , as discussed herein . Sequences In certain embodiments , oligomers as long as 40 bases 45 may be suitable , where at least a minimum number of bases , Target Targeting TS SEQ e . g ., 10 - 12 bases , are complementary to the target sequence . Gene Sequence ( TS ) * ID NO : In general, however , facilitated or active uptake in cells is optimized at oligomer lengths of less than about 30 or less ???? CTTCGATAGTG than about 20 bases. Included are antisense oligomers that ???? ATATCGCTCAC N consist of about 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21, 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 , 31, 32 , 33 , 34 , 35 , ???? ?????cc???? w

36 , 37 , 38 , 39 , or 40 bases , in which at least about 6 , 8 , 9 , A 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , ???? CACAGGAATTC 26 , 27 , 28 , 29, 30 , 31, 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , or 40 acps TTGCCATTAGC o contiguous or non - contiguous bases are complementary to a 55 target gene described herein , for example , a target sequence ??? - ? CTGTAGTGATTTCACCA o of Table 1 ( e . g . , SEQ ID NO : 69 or 70 ) . In certain embodiments , antisense oligomers may be fabA TTATCTACCAT 100 % complementary to the target sequence , or may include fabB GCACGTTTCAT o mismatches, e . g . , to accommodate variants , as long as a 60 fabI heteroduplex formed between the oligomer and target AGAAAACCCAT o sequence is sufficiently stable to withstand the action of cellular nucleases and other modes of degradation which gapA TTGATAGTCAT may occur in vivo , and reduce expression of the targeted accA GCTTTTTTCAT t mRNA . Hence , certain oligomers may have about or at least 65 * The thymines ( T ) can be uracils ( U ) , and vice versa ; I is about 70 % sequence complementarity, e . g ., 70 % , 71 % , inosine , 72 % , 73 % , 74 % , 75 % , 76 % , 77 % , 78 % , 79 % , 80 % , 81 % , US 10, 391, 098 B2 21 22 TABLE 2B TABLE 2B - continued Exemplary targeting sequences Exemplary targeting sequences associated with other biochemical associated with other biochemical pathways and / or cellular processes pathways and / or cellular processes Target Targeting TS SEQ Target Targeting TS SEQ Gene Sequence ( TS ) * ID NO : Gene Sequence ( TS ) * ID NO : mura ATCCATTTAGT 12 boxA CTCTTAATGAT 49 mura CATTTAGTTTG 13 10 boxc ATCCACACAAG 50 murA AATTTATCCAT 14 rpoD - E TCCACCAAGTCACCA 51 mura AAATTTATCCA 15 pryc AGAGTTCAAGG 52 rpmB ACTCGGGACAT 16 cara GGTGCTCAAAC 53 rpmB ?????c????? adeA ATACTGTCCAA 54 rpmB GGCAGACTCGG 18 blaT ?????ccTTTT 55 rpmB CTTAGACATGG 19 20 cml TCCTTCTGATT 56 adk ATGATACGCAT 20 * The thymines ( T ) can be uracils ( U ) , and vice versa ; I is infA TCTTTGGCCAT 21 inosine , Certain antisense oligomers thus comprise , consist , or ftsz TCAAATGAGGC 22 25 consist essentially of a targeting sequence in Tables 2A - B ftsz AATGAGGCCAT 23 ( e . g ., SEQ ID NOS: 1 - 56 ) or a variant or contiguous or non - contiguous portion ( s ) thereof. For instance , certain anti ftsz ATAGTTTCTCTCC sense oligomers comprise about or at least about 6 , 7 , 8 , 9, rpoD TCATCTTTGCT 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21, 22 , 23 , 24 , 25 , 30 26 , or 27 contiguous or non - contiguous nucleotides of any rpoD TTTTGCTCCAT 26 of the targeting sequences in Tables 2A - B ( e . g . , SEQ ID aroc TTCCCTGCCAT 27 NOS: 1 - 56 ) . For non - contiguous portions, intervening nucleotides can be deleted or substituted with a different aroc TTTCCAGCCAT 28 nucleotide , or intervening nucleotides can be added . Addi 35 tional examples of variants include oligomers having about murF ACGCTAATCAT 29 or at least about 70 % sequence identity or homology , e . g . , 1pxC TGTTTGATCAT 30 70 % , 71 % , 72 % , 73 % , 74 % , 75 % , 76 % , 77 % , 78 % , 79 % , 80 % , 81 % , 82 % , 83 % , 84 % , 85 % , 86 % , 87 % , 88 % , 89 % , kata AATTCGAGCAT 31 90 % , 91 % 92 % , 93 % , 94 % , 95 % , 96 % , 97 % , 98 % , 99 % or 40 100 % sequence identity or homology , over the entire length boxA TGTTAAAGAGC of any of the targeting sequences in Tables 2A - B (e . g ., SEQ rpoD - E CTTGTAACCACACCA 33 ID NOS : 1 - 56 ) . The activity of antisense oligomers and variants thereof pryc GGTGCAGTCAT 34 can be assayed according to routine techniques in the art pryA GACTTAATCAA 45 ( see , e . g ., the Examples ) . III . Antisense Oligomer Compounds lgt CTACTGGTCAT 36 The antisense oligomers typically comprises a base fola CATTGAGATTT sequence of sufficient length and complementarity to spe cifically hybridize to a bacterial mRNA target sequence that infB ACATCTGTCAT 38 50 encodes a protein associated with a biochemical pathway and /or cellular process, and thereby reduce expression ( e .g ., nrda TTCTGATTCAT 3 translation ) of the protein . This requirement is optionally nrdB GTATATGCCAT 40 met when the oligomer compound has the ability to be actively taken up by bacterial cells , and once taken up , form zipA TCCTGCATCAT 41 55 a stable duplex ( or heteroduplex ) with the target mRNA , optionally with a Tm greater than about 40° C . or 45° C . coaA ATATACCTCAT A . Antisense Oligomer Chemical Features gyrA - E GTTACCCTGACCGACCA 3 In certain embodiments , the backbone of the antisense oligomer is substantially uncharged , and is optionally rec gyrA - E GTTACCCTGACCACCA 44 60 ognized as a substrate for active or facilitated transport mrda TGTTTCATACG 45 across a cell wall and / or cell membrane . The ability of the oligomer to form a stable duplex with the target RNA may 1pxB GGTTTGCCAAG 46 also relate to other features of the backbone , including the 1pxc TGTTTCACCAT 47 length and degree of complementarity of the antisense 65 oligomer with respect to the target, the ratio of G : C to A : T kata TTTTTCGCCAA 48 base matches, and the positions of any mismatched bases. The ability of the antisense oligomer to resist cellular US 10 , 391, 098 B2 23 24 nucleases may promote survival and ultimate delivery of the base in a polynucleotide . Y or V2 may be oxygen , sulfur, agent to the cell. Exemplary antisense oligomer targeting nitrogen , or carbon , preferably oxygen . The X moiety pen sequences are listed in Tables 2A - B ( supra ) . dant from the phosphorus may be fluorine , an alkyl or In certain embodiments , the antisense oligomer is a mor substituted alkyl, an alkoxy or substituted alkoxy , a thio pholino -based oligomer , for example , a phosphorodiamidate 5 alkoxy or substituted thioalkoxy, or unsubstituted , mono morpholino oligomer (PMO ). Morpholino -based oligomers substituted , or disubstituted nitrogen , including cyclic struc refer to an oligomer comprising morpholino subunits sup - tures, such as morpholines or piperidines . Alkyl, alkoxy and porting a nucleobase and , instead of a ribose, contains a thioalkoxy include 1 -6 carbon atoms. The Z moieties may be morpholine ring. Exemplary internucleoside linkages sulfur or oxygen , and are preferably oxygen . include , for example , phosphoramidate or phosphorodiami- 1 Accordingly , various embodiments of the disclosure date internucleoside linkages joining the morpholine ring include a substantially uncharged antisense morpholino oli nitrogen of one morpholino subunit to the 4 ' exocyclic gomer, composed of morpholino subunits and phosphorus carbon of an adjacentmorpholino subunit . Each morpholino containing intersubunit linkages joining a morpholino nitro subunit comprises a purine or pyrimidine nucleobase effec - 15 gen of one subunit to a 5 ' - exocyclic carbon of an adjacent tive to bind , by base - specific hydrogen bonding , to a base in subunit, and having (a ) about 10 -40 nucleotide bases , and an oligomer. ( b ) a targeting sequence of sufficient length and comple Morpholino - based oligomers ( including antisense oli mentarity to specifically hybridize to a bacterial mRNA gomers ) are detailed , for example , in U .S . Pat. Nos. 5 ,698 , target sequence that encodes a protein associated with a 685 ; 5 ,217 , 866 ; 5 , 142, 047 ; 5 , 034 , 506 ; 5 , 166 ,315 ; 5 , 185 , biochemical pathway and / or cellular process , or a protein 444 ; 5 ,521 , 063; 5 , 506 ,337 and pending U . S . patent associated with antibiotic resistance, as described herein . In application Ser. Nos. 12 /271 , 036 ; 12/ 271 ,040 ; and PCT some instances, the oligomer is conjugated to a cell - pen Publication No . WO /2009 /064471 and WO /2012 /043730 etrating peptide (CPP ) . and Summerton et al. 1997 , Antisense and Nucleic Acid In various aspects , an antisense oligomer of the disclosure Drug Development, 7 , 187 - 195 , which are hereby incorpo - os includes a compound of formula ( 1 ) rated by reference in their entirety . Within the oligomer structure , the phosphate groups are commonly referred to as forming the “ internucleoside link ages ” of the oligomer. The naturally occurring internucleo side linkage of RNA and DNA is a 3 ' to 5 ' phosphodiester 2 linkage . A “ phosphoramidate ” group comprises phosphorus 30 __ Nu having three attached oxygen atoms and one attached nitro gen atom , while a " phosphorodiamidate ” group comprises phosphorus having two attached oxygen atoms and two attached nitrogen atoms. In the uncharged or the cationic z P - R internucleoside linkages of the morpholino -based oligomers ZA described herein , one nitrogen is always pendant to the 0 linkage chain . The second nitrogen , in a phosphorodiami date linkage , is typically the ring nitrogen in a morpholine TONN7 ring structure . In particular embodiments , the morpholino subunits are 40* joined by phosphorous- containing intersubunit linkages in accordance with the structure : O = P - R1

Z = P1 - X La Nu

R2 R3 or a pharmaceutically acceptable salt thereof, where each Nu is a nucleobase which taken together 55 forms a targeting sequence ; where Y , = oxygen ( 0 ) or sulfur, nitrogen , or carbon ; X is an integer from 9 to 38 ; Z = oxygen or sulfur, preferably oxygen ; Pj is a purine or T is selected from OH and a moiety of the formula : pyrimidine base -pairing moiety effective to bind , by base specific hydrogen bonding , to a base in a polynucleotide , and X is — NRR ' where R and R ' are the same or different 60 Ro and are either H or alkyl. In particular embodiments , X is - NRR ' , where R and R ' are the same or different and are O = — N (R4 ) 2R5 , either H or methyl. Also included are antisense oligomer that comprise a é sequence of nucleotides of the formula in FIGS . 1A - 1E . In 65 FIG . 1A , B is a purine or pyrimidine base -pairing moiety effective to bind , by base -specific hydrogen bonding , to a US 10 , 391 , 098 B2 25 26 where each R4 is independently C . -C . alkyl, and R is In certain embodiments , T is selected from : selected from an electron pair and H , and Róis selected from OH , — N (R7 ) CH C ( O )NH2 , and a moiety of the formula : s HOTHot NH2 N - R8 ,

10 where : 0 = — N ( CH3) 2 ; O = — N ( CH3) 2 ; R ? is selected from H and C , - C alkyl, and R $ is selected from G , C ( O ) R 'OH , acyl, trityl, and 4 -methoxytrityl , where : Rº is of the formula ( O - alkyl) , , - wherein y is an 15 nmn integer from 3 to 10 and each of the y alkyl groups is independently selected from C2- C6 alkyl; each instance of Rl is — N (R1 ) R11 wherein each R10 is independently C , -C6 alkyl, and R " is OH selected from an electron pair and H ; R2 is selected from H , G , acyl, trityl, 4 -methoxytrityl , ma n ; and O = P - N (CH3 ) . benzoyl, stearoyl, and a moiety of the formula : 25 mi In some embodiments , R2 is selected from H , G , acyl , trityl, 4 -methoxytrityl , benzoyl, and stearoyl. In various embodiments, T is selected from : 30 Hot - oj O N 35 ONH (R12 ) N N N ( R12 )2 , where L is selected from - C (O )( CH2 ) 6C ( 0 ) and 40 O = P - N (CH3 )2 ; O = P - N (CH3 ) 2 ; and C ( O ) (CH2 ) 2S , (CH2 ) 2C ( O ) - , and each R12 is of the for mula (CH2 ) 2OC ( O ) N ( R ! 4 ) 2 wherein each R14 is of the formula ( CH2) 6NHC ( NH ) NH2; and R® is selected from an electron pair , H , and C1- C6 alkyl, OH wherein G is a cell penetrating peptide (" CPP ” ) and linker 45 moiety selected from - C ( O ) (CH2 ) sNH -CPP , C ( O ) ( CH2 )- NH- CPP, C ( O )( CH2 )- NHC( O )( CH2 ) NH- CPP , and R2 is G . and C ( O )CH NH -CPP , or G is of the formula : In some embodiments , T is of the formula : 50

0 = — N (CH3 ) 2 , 55 wherein the CPP is attached to the linker moiety by an amide bond at the CPP carboxy terminus, with the proviso that only one instance of G is present, 60. Róis of the formula : wherein the targeting sequence specifically hybridizes to a bacterial mRNA target sequence that encodes a protein associated with a biochemical pathway and /or cellular pro cess . In some embodiments , X is from 9 to 18 . In certain 65 R 'OH , embodiments , X is 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , man 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 or 30 . and R2 is G . US 10 , 391 ,098 B2 27 28 In certain embodiments , T is of the formula :

Hot to Nu

O = P - N (CH3 ) 2 UZ — N (CH3 ) 2 , La Nu and R² is G . O = P - N ( CH3) 2 In certain embodiments , T is of the formula: 0 Nu

0-Z

R2 R3 or a pharmaceutically acceptable salt thereof, O = P - N (CH3 ) 2 . 30 where each Nu is a nucleobase which taken together forms a targeting sequence ; X is an integer from 9 to 28 ; T is selected from : 35 In some embodiments , R² is G or T is of the formula : Hotty NH2 40

N Z

O = P - N (CH3 ) 2 ; O = -P - N (CH3 ) 2 ; O = P 45

50 In some embodiments , R² is selected from H , acyl, trityl, 4 -methoxytrityl , benzoyl, and stearoyl. In various embodiments , R2 is selected from H or G , and 55 R3 is selected from an electron pair or H . In a particular embodiment , R2 is G . In some embodiments, R² is H or acyl. In some embodiments , each R ' is N (CH3 ) 2. In some OH N embodiments , at least one instance of R ' is — N (CH3 ) 2 . In m ini and O = P - N ; certain embodiments , each instance of Rl is N (CH3 ) . In various embodiments of the disclosure , an antisense oligomer of the disclosure includes a compound of formulas ( II ): US 10 ,391 ,098 B2 29 30 R2 is selected from H , G , acyl, trityl, 4 -methoxytrityl , benzoyl, and stearoyl; and (III )

R® is selected from an electron pair, H , and C -C6 alkyl, 0 Nu wherein G is a cell penetrating peptide (“ CPP” ) and linker moiety selected from - C (O )( CH2 ) 3NH -CPP , C (O ) (CH2 ) 2NH - CPP, C ( O ) (CH2 ) 2NHC ( O )( CH2) 5NH -CPP , O = P - R and - C ( O ) CH NH - CPP, or G is of the formula : 10

La Nu

O = P - R

Jx - o . Nu wherein the CPP is attached to the linker moiety by an amide bond at the CPP carboxy terminus, with the proviso that only 25 one instance of G is present. In some embodiments , T is G R2 TEG as defined above, R2 is G , and R3 is an electron pair or H . In certain embodiments , R² is selected from H , acyl, or a pharmaceutically acceptable salt thereof, where each Nu is a nucleobase which taken together trityl, 4 -methoxytrityl , benzoyl , and stearoyl and T is of the forms a targeting sequence ; formula : X is an integer from 9 to 28 ; T is selected from : 35 - Hotho

/ O = P - N ( CH3) 2 ; 45

NH2 In some embodiments , R2 is G or T is of the formula : 50 R9 OH Z-AGOP - N ( CH3) 2 ; and w inn ; 55

each instance of R ' is — N ( R19) , R11 wherein each R10 is 60 independently C . - C . alkyl, and R1 is selected from an electron pair and H ; R2 is selected from an electron pair, H , and C , - Co alkyl; and 65 G is a cell penetrating peptide ( “ CPP ” ) and linker moiety In various aspects, an antisense oligomer of the disclosure selected from C (O ) (CH2 ) -NH -CPP , - C (O ) (CH2 ) 2NH includes a compound of formula ( III) : CPP, C ( O )( CH2 )- NHC( O )( CH ) NH- CPP, US 10 , 391 ,098 B2 31 32 and C (O )CH NH -CPP , or G is of the formula : In various aspects , an antisense oligomer of the disclosure can be a compound of formula ( V ) :

0-2

wherein the CPP is attached to the linker moiety by an amide 10 Z-e bond at the CPP carboxy terminus. In some embodiments , at O = P - RI least one instance of Rl is N (CH3 ) 2 . In certain embodi 0 ments, each instance of R1 is — N (CH2 ) , O Nu In various aspects , an antisense oligomer of the disclosure includes a compound of formula (IV ) : O = P - RI Hotto

N -A - O = P - R O = P - R -0 -

0 Nu obtbra Nu

O = -2P - R -0 LO. Nu wherein : X is an integer from 9 to 18 ; 35 U-Z each Nu is a nucleobase which taken together forms a = P - R - targeting sequence ; 0 each instance of R ' is - N (R19 ), R11 wherein each R10 is 40 independently CZ- C6 alkyl, and Rll is selected from an Z electron pair and H ; -0 R² is selected from H , trityl, 4 -methoxytrityl , acyl, ben zoyl, and stearoyl; and or a pharmaceutically acceptable salt thereof, wherein : 45 X is an integer from 9 to 28 ; R3 is selected from an electron pair , H , and C . - C , alkyl, each Nu is a nucleobase which taken together forms a wherein G is a cell penetrating peptide (“ CPP ” ) and linker targeting sequence ; each instance of R ' is N ( R ! ' ), R11 wherein each R10 is moiety selected from _ C ( O ) (CH2 ) - NH -CPP , - C ( O ) independently CZ - C6 alkyl, and Rll is selected from an (CH2 ) 2NH -CPP , - C ( O ) (CH2 ) 2NHC ( O ) (CH2 ) 3NH -CPP , electron pair and H ; and and - C (O ) CH NH - CPP , or G is of the formula : G is a cell penetrating peptide (“ CPP ” ) and linker moiety selected from C ( O ) (CH2 ) -NH -CPP , C ( O ) (CH2 ) 2NH CPP, C ( O )( CH2 )- NHC( O )( CH2 ) - NH- CPP, and C ( O ) CH NH - CPP, or G is of the formula : 55

wherein the CPP is attached to the linker moiety by an amide bond at the CPP carboxy terminus . In some embodiments , at least one instance of R ' is - N (CH3 ) 2. In certain embodi wherein the CPP is attached to the linker moiety by an amide ments , each instance of R ' is N (CH2 ) , . bond at the CPP carboxy terminus. In some embodiments , at 65 least one instance of Rl is N (CH2 ) . In certain embodi- In various aspects , an antisense oligomer of the disclosure ments , each instance ofR is - N (CH3 )2 . includes a compound of formula (VI ) : US 10 ,391 ,098 B2 34 33 TABLE C1 Exemplary Cell - Penetrating Peptides Z-9

SEO Name Sequence ID NO : O = PN - N (CH3 ) 2 (RXR ) 4 RXRRXRRXRRXR 57 ( RFF ) 3R RFFRFFRFFR 58 Nu 10 ( RXR ) 4XB RXRRXRRXRRXRXB 59

Z-A O = P - N (CH3 ) 2 ( RFF ) 3RXB RFFRFFRFFRXB 60 -0 (RFR ) 4 RFRRFRRFRRFR 61 Nu (RYR ) 4 RYRRYRRYRRYR 62

2 ( RGR ) 4 RGRRGRRGRRGR 63 O = P - N (CH3 ) 2 20 ( RFR ) 4XB RFRRFRRFRRFRXB 64

Nu ( RYR ) 4XB RYRRYRRYRRYRXB 65

( RGR ) 4XB RGRRGRRGRRGRXB 66 25 ( RFF ) 3RXB RFFRFFRFFRXB 67 or a pharmaceutically acceptable salt thereof, wherein : (RFF ) 3RG RFFRFFRFFRG 30 x is 6 - aminohexanoic acid ; Bis B - alanine ; F is phenylalanine ; X is an integer from 9 to 28 ; Y is tyrosine ; G is glycine ; R is arginine each Nu is a nucleobase which taken together forms a targeting sequence ; In some embodiments , the CPP is linked at its C - terminus to the 3 '- end or the 5 ' - end of the oligomer via a 1 , 2 , 3 , 4 , or R² is selected from H or acyl; and 5 amino acid linker . Gis a cell penetrating peptide (“ CPP ” ) and linker moiety 35 CPPs, their synthesis , and methods of conjugating a CPP selected from - C ( O ) ( CH2) - NH - CPP , C ( O ) ( CH2) 2NH to an oligomer are detailed , for example , in International CPP, C ( O )( CH3 )2NHC ( O ) ( CH2) NH- CPP, Patent Application Publication Nos. WO 2004 /097017 , WO and - C ( O )CH NH -CPP , or G is of the formula : 2009 /005793 , and WO 2012 / 150960, which are all incor 40 porated by reference in their entirety . In some embodiments , the CPP is linked at its C - terminus to the 3' - end or the 5' - end of the oligomer via a 1, 2 , 3 , 4 , or 5 amino acid linker . In particular embodiments , including antisense oligomer compounds of formula ( 1 ) - (VI ) , the link 45 ers can include : - C ( O )( CH2) ; NH - CPP ( X linker ), C ( O ) ( CH2) 2NH -CPP ( B linker ), C ( O ) (CH2 ) 2NHC ( O ) ( CH2) -NH - CPP (XB peptide linker) , and C (O )CH NH wherein the CPP is attached to the linker moiety by an amide C PP ( G linker ), or formula : bond at the CPP carboxy terminus. The antisense oligomers can be prepared by stepwise 50 solid -phase synthesis , employing methods known in the art CPP, and described in the references cited herein . B B . Cell- Penetrating Peptides In certain embodiments , the antisense oligomer is conju - * gated to a cell -penetrating peptide (CPP ) . In some embodi ments , the CPP is an arginine -rich peptide . By “ arginine -rich carrier peptide” is meant that the CPP has at least 2 , and wherein the CPP is attached to the linker moiety by an amide preferably 2 , 3 , 4 , 5 , 6 , 7 , or 8 arginine residues , each 60 bond at the CPP carboxy terminus. In some embodiments of optionally separated by one or more uncharged , hydropho the disclosure , including antisense oligomer compounds of bic residues, and optionally containing about 6 - 14 amino formula ( 1 ) - (VI ) , G is selected from SEQ ID NOS : 59, 60 , acid residues . FIGS. 1F - 1H show exemplary chemical struc - and 64 -68 . In various embodiments , including antisense tures of CPP - PMO conjugates used in the Examples , includ - oligomer compounds of formula (1 ) - (VI ) , the CPP is selected ing 5 ' and 3 ' PMO conjugates . 65 Tromfrom SASEQ ID NO : 57 , and 61 -63 . Exemplary CPPs are provided in Table C1 (SEQ ID In some embodiments , including antisense oligomer com NOS: 57 -68 ) . pounds of formula ( I) - (VI ) , the CPP is selected from : US 10 , 391 , 098 B2 35 36 -continued Ra; N Ra ;

NH HN HN NH2 HN ANH NH NH | L HNCNH , H?N NH _ and H2NNH

HN .

Ntro: 25

HN NH

H?N SNH HN ANH

ER ,

HN NH2

HetNH " ?? 50 wherein Ra is selected from H , acetyl, benzoyl, and stearoyl. HN ” NH2H2NNH In some embodiments , including antisense oligomer com pounds of formula (I ) - ( VI) , G is selected from :

N - Ra ; NH

NH HN HNNH , H2NNH US 10 , 391 , 098 B2 31 38 -continued H2NNH

N N

HEN

H , N . NH

HN .

N - RQ.

HNNH

| Pa, N

* NH NH

HN NH , HN ANH

- Ra; and N

NH NH HNCNH , H?N NHÉ US 10 ,391 , 098 B2 40 39 -continued NH

H N NH

HN NH

wherein Ra is selected from H , acetyl , benzoyl, and stearoyl. In various aspects , an antisense oligomer of the disclo - sure, or a pharmaceutically acceptable salt thereof, includes an antisense oligomer of the formula (VII ) selected from : US 10 ,391 ,098 B2 41

(VIIA) VIIB)( ? ZE

IZ HN ????“? Nu

NH ????? (H3C)2N Nu

N(CH3)2

N(CH3)2 intanyangtingnytNu .01 11

(H3C)2N Nu

N(CH3)2

5 Nu N(CH3)2o fryezoterapijeTommyNH H2NNH

N HN HN?NH REN. US 10, 391 , 098 B2 43 44

(VIIC) (VIID) `NER,

SR6; H2NNH HN ZI N

}

(H3CN) Nu HN H2NNH N(CH3)2

O Nu . ???????-???????? ??-Lov???????5.N(CH3)20 -continued x

N H3C()2N Nu

N(CH2)2 NH Nu HNNH2

3 N(CH2)2

Z IZ *NH HNNH RL ?- US 10, 391 , 098 B2 45 ,

(VIIE) (VIIF)

HN HN. 1

3 Nu 1 TZ 1

(HC)2N Nu

HN HNNH NCH(): Nu

{ -- continued- N(CH3)2,

(HC)2N Nu

N(CH3), HNNH2 NH N Nu

N(CH3)2

CNH,

N NH HN

HO. -?????????????? R US 10, 391 , 098 B2 LV

(VIIG) (VIIH) -R; NSNHT. NH

N ) NH HNTNH, 07 N

(MI,C2N Nu

yrad,> N(CH3)2

N(CH3)2

-continued Nu

(H3C)2N Nu

N(CH3)2 --S-???…???/

N Nu

N(CH3)2O NH HN. H?N

IZ HNNH, HN.

HO R T US 10 ,391 ,098 B2 49 50

(VIII) Ra; (VIIJ) H2NNH. *?? RDE

3' Nu i

.] NH HN?NH HN (H3C)N HO. Nu

N(CH3)|

Nu ZI NICIN(CH3)2o

-continued 3' Nu

X - Bisfemintapetotpet

(H3C)2N Nil Nu

N(CH3)2

OH14

N(CH3)2o^ HNNH NH

NH HNANH, HN. mootopónimponente ?Ra- SI US 10, 391 , 098 B2

(VIIK) -and (VIIL)

NH2 NH HNCNH ? NH Nu ) HN?

1 -

A Last (HCAN Nu

CH3)2N(

I Á

??????“?y??-?????.N(CH3)201 Nu -continued , Xr

(H3C)2N Nu

N(CH3)2

N(CH3)2o

} NH NH2 HN. NH H?N, HN

Ro+? OH US 10 , 391 , 098 B2 53 54 wherein X is an integer from 9 to 38 , Rº is selected from H , In certain embodiments , the protein associated with a acetyl, benzoyl, and stearoyl, R ' is selected from H , acetyl, biochemical pathway and/ or cellular process is an RNA benzoyl, stearoyl, trityl, and 4 -methoxytrityl , and each Nu is synthesis protein . In some embodiments , the RNA synthesis a purine or pyrimidine base -pairing moiety which taken protein is a sigma D factor of RNA polymerase . For together form a targeting sequence described above . 5 example , in certain embodiments , the sigma D factor of C . Antisense Oligomer Targeting Sequences RNA polymerase is RpoD . In various embodiments of the antisense oligomers of the In some embodiments , the protein associated with a disclosure , including the antisense oligomer compounds of biochemical pathway and /or cellular process is an aromatic formulas ( 1 ) - (VII ) , the targeting sequence can specifically compound biosynthesis protein . In certain embodiments , the hybridizes to a bacterial mRNA target sequence that encodes 10 aromatic compound biosynthesis protein is a chorismate a protein associated with a biochemical pathway and/ or synthase ( 5 - enolpyruvylshikimate - 3 -phosphate phospho cellular process , or a protein associated with antibiotic lyase ) . For example, in some embodiments , the chorismate resistance . In some embodiments , the target sequence com - synthase ( 5 - enolpyruvylshikimate - 3 - phosphate phospho prises a translational start codon of the bacterial mRNA lyase ) is AroC . and / or a sequence within about 30 bases upstream or down - 15 In some embodiments , the protein associated with anti stream of the translational start codon of the bacterial biotic resistance is selected from one or more of BlaT, Cml, mRNA . and AdeA . In various embodiments , the protein associated with a In some embodiments where the protein associated with biochemical pathway and / or cellular process may be a fatty a biochemical pathway and /or cellular process may be a acid biosynthesis protein . In some embodiments , the fatty 20 peptidoglycan biosynthesis protein , a ribosomal protein , a acid biosynthesis protein can be an acyl carrier protein . In cellular energy homeostasis protein , a protein biosynthesis certain embodiments , the acyl carrier protein may be Acpp . protein , a cell division protein , an RNA synthesis protein , an In some embodiments , the fatty acid biosynthesis protein aromatic compound biosynthesis protein , or antibiotic resis may be a carboxyltransferase alpha subunit of an acetyl tance , the targeting sequence comprises or consists of at Coenzyme A carboxylase . In certain embodiments, the car - 25 least one of the targeting sequences set forth in Table 2B boxyltransferase alpha subunit of an acetyl Coenzyme A ( e . g . , SEQ ID NOS : 12 - 56 ) , comprises or consists of a carboxylase may be AccA . In some embodiments , the target fragment of at least 10 contiguous nucleotides of a targeting sequence may be SEQ ID NOs: 1 - 11, wherein thymine bases sequence in Table 2B ( e . g ., SEQ ID NOS : 12 - 56 ) , or ( T ) are optionally uracil bases ( U ) . In certain embodiments, comprises or consists of a variant having at least 80 % the targeting sequence comprises or consists of at least one 30 sequence identity to a targeting sequence in Table 2B ( e . g . , of the targeting sequences in Table 2A ( e . g . , SEQ ID NOS: SEQ ID NOS : 12 - 56 ) , wherein thymine bases ( T ) are 1 - 11 ) , comprises or consists of a fragment of at least 10 optionally uracil bases ( U ) . contiguous nucleotides of a targeting sequence in Table 2A In certain embodiments , including the antisense oligomer ( e . g ., SEQ ID NOS: 1 - 11) , or comprises or consists of a compounds of formulas ( 1 ) - ( VII ), the targeting sequence is variant having at least 80 % sequence identity to a targeting 35 selected from : sequence in Table 2A ( e . g . , SEQ ID NOS: 1 - 11 ) , wherein a ) SEQ ID NO : 1 (CTTCGATAGTG ) wherein X is 9 ; thymine bases ( T ) are optionally uracil bases ( U ) . b ) SEO ID NO : 2 (ATATCGCTCAC ) wherein X is 9 ; In some embodiments , the protein associated with a c ) SEQ ID NO : 3 (ATTCTCCTCAT ) wherein X is 9 ; biochemical pathway and / or cellular process may be a d ) SEQ ID NO : 4 (CACAGGAATTC ) wherein X is 9 ; peptidoglycan biosynthesis protein . In certain embodiments , 40 e ) SEQ ID NO : 5 ( TTGCCATTAGC ) wherein X is 9 ; the peptidoglycan biosynthesis protein can be a UDP- N - f ) SEQ ID NO : 6 (CTGTAGTGATTTCACCA ) wherein acetylglucosamine 1 - carboxyvinyltransferase . In some X is 15 ; embodiments , the UDP- N -acetylglucosamine 1 - carboxyvi g ) SEQ ID NO : 7 ( TTATCTACCAT) wherein X is 9 ; nyltransferase may be MurA . h ) SEO ID NO : 8 (GCACGTTTCAT ) wherein X is 9 ; In some embodiments, the protein associated with a 45 i ) SEQ ID NO : 9 ( AGAAAACCCAT ) wherein X is 9 ; biochemical pathway and / or cellular process is a ribosomal j ) SEQ ID NO : 10 ( TTGATAGTCAT ) wherein X is 9 ; and protein . In certain embodiments , the ribosomal protein is a k ) SEQ ID NO : 11 (GCTTTTTTCAT ) wherein X is 9 , 50S ribosomal protein L28 . In some embodiments , the 50S wherein thymine bases ( T ) may be uracil bases ( U ) . ribosomal protein L28 is RpmB. In some embodiments , including the antisense oligomer In various embodiments , the protein associated with a 50 compounds of formulas ( 1) - (VII ) , the targeting sequence is biochemical pathway and /or cellular process is a cellular selected from : energy homeostasis protein . In some embodiments , the a ) SEQ ID NO : 12 (ATCCATTTAGT ) wherein X is 9 ; cellular energy homeostasis protein is an adenylate kinase . b ) SEQ ID NO : 13 (CATTTAGTTTG ) wherein X is 9 ; In certain embodiments , the adenylate kinase is Adk . c ) SEQ ID NO : 14 (AATTTATCCAT ) wherein X is 9 ; In some embodiments , the protein associated with a 55 d ) SEO ID NO : 15 (AAATTTATCCA ) wherein X is 9 ; biochemical pathway and/ or cellular process is a protein e ) SEQ ID NO : 16 ( ACTCGGGACAT ) wherein X is 9 ; biosynthesis protein . In certain embodiments , the protein f ) SEO ID NO : 17 (CTATTCTCCAA ) wherein X is 9 ; biosynthesis protein is a translation initiation factor . In g ) SEQ ID NO : 18 (GGCAGACTCGG ) wherein X is 9 ; various embodiments , the translation initiation factor is h ) SEQ ID NO : 19 (CTTAGACATGG ) wherein X is 9 ; InfA . 60 i ) SEQ ID NO : 20 ( ATGATACGCAT ) wherein X is 9 ; In some embodiments , the protein associated with a j ) SEQ ID NO : 21 ( TCTTTGGCCAT ) wherein X is 9 ; biochemical pathway and /or cellular process is a cell divi- k ) SEQ ID NO : 22 (TCAAATGAGGC ) wherein X is 9 ; sion protein . In certain embodiments , the cell division 1 ) SEQ ID NO : 23 ( AATGAGGCCAT) wherein X is 9 ; protein is a protein that assembles into a ring at the future m ) SEQ ID NO : 24 ( ATAGTTTCTCTCC ) wherein X is site of the septum of bacterial cell division . For example , in 65 11 ; some embodiments , the protein that assembles into a ring at n ) SEQ ID NO : 25 ( TCATCTTTGCT) wherein X is 9 ; the future site of the septum of bacterial cell division is FtsZ 0 ) SEQ ID NO : 26 (TTTTGCTCCAT ) wherein X is 9 ; US 10 , 391 ,098 B2 55 56 p ) SEQ ID NO : 27 (TTCCCTGCCAT ) wherein X is 9 ; hh ) SEQ ID NO : 45 ( TGTTTCATACG ) wherein X is 9 ; q ) SEQ ID NO : 28 (TTTCCAGCCAT ) wherein X is 9 ; ii ) SEQ ID NO : 46 (GGTTTGCCAAG ) wherein X is 9 ; r ) SEQ ID NO : 29 (ACGCTAATCAT ) wherein X is 9 ; jj ) SEQ ID NO : 47 ( TGTTTCACCAT ) wherein X is 9 ; s ) SEQ ID NO : 30 ( TGTTTGATCAT) wherein X is 9 ; kk ) SEO ID NO : 48 ( IIIITCGCCAA ) wherein X is 9 ; 11 ) SEO ID NO : 49 (CTCTTAATGAT ) wherein X is 9 ; t ) SEQ ID NO : 31 (AATTCGAGCAT ) wherein X is 9 ; 5 mm ) SEQ ID NO : 50 ( ATCCACACAAG ) wherein X is 9 ; u ) SEQ ID NO : 32 ( TGTTAAAGAGC ) wherein X is 9 ; nn ) SEQ ID NO : 51 ( TCCACCAAGTCACCA ) wherein V ) SEQ ID NO : 33 ( CTTGTAACCACACCA ) wherein X X is 13 ; is 13 ; 00 ) SEQ ID NO : 52 (AGAGTTCAAGG ) wherein X is 9 ; w ) SEQ ID NO : 34 (GGTGCAGTCAT ) wherein X is 9 ; o pp ) SEQ ID NO : 53 (GGTGCTCAAAC ) wherein X is 9 , x ) SEQ ID NO : 35 GACTTAATCAA( ) wherein X is 9 ; 10 wherein thymine bases ( T ) may be uracil bases ( U ) . y ) SEQ ID NO : 36 (CTACTGGTCAT ) wherein X is 9 ; In some embodiments of the disclosure , including the Z ) SEQ ID NO : 37 (CATTGAGATTT ) wherein X is 9 ; antisense oligomer compounds of formulas (I ) -( VII ) , the aa ) SEQ ID NO : 38 (ACATCTGTCAT ) wherein X is 9 ; targeting sequence is selected from : bb ) SEO ID NO : 39 (TTCTGATTCAT ) wherein X is 9 ; 10 a ) SEQ ID NO : 54 ( ATACTGTCCAA ) ; cc ) SEQ ID NO : 40 (GTATATGCCAT ) wherein X is 9 ; b ) SEQ ID NO : 55 (CTCTTCCTTTT ) ; and dd ) SEO ID NO : 41 ( TCCTGCATCAT ) wherein X is 9 ; SEO ID NO : 56 ( TCCTTCTGATT) , ee ) SEQ ID NO : 42 ( ATATACCTCAT ) wherein X is 9 ; wherein X is 9 , and thymine bases ( T ) may be uracil bases ff ) SEQ ID NO : 43 (GTTACCCTGACCGACCA ) (U ). wherein X is 15 ; o D . Exemplary Antisense Oligomers gg ) SEQ ID NO : 44 (GTTACCCTGACCACCA ) wherein 20 Exemplary antisense oligomers (AONs ) of the disclosure X is 14 ; include those described in Tables 3A - B below . TABLE 3A Exemplary Fatty Acid Biosynthesis - Associated Targeting Sequences AONS Targeting 5 ! 31 PMO Target Sequence TS SEQ Attachment Attachment CPP SEQ Name Gene ( TS ) * ID NO : * * * * * ID NO .

PPMO # 1 acpP CTTCGATAGTG - ( RXR ) 4XB - Acetyl 59 PPMO # 2 acpp CTTCGATAGTG 1 TEG ( RXR ) 4XB 59

PPMO # 3 acpP CTTCGATAGTG - ( RFR ) 4XB Acetyl PPMO # 4 acpP CTTCGATAGTG 1 TEG ( RYR ) 4XB 59 PPMO # 5 acpp CTTCGATAGTG 1 TEG ( RGR ) 4XB

PPMO # 6 acpP ATATCGCTCAC N ( RXR ) 4XB - Acetyl 59 PPMO # 7 acpp ATATCGCTCAC 2 TEG ( RXR ) 4XB 59

PPMO # 8 acpP ?????cc???? N (RXR ) 4XB Acetyl 59

PPMO # 9 acpP ATATCGCTCAC N (RFR ) 4XB Acetyl 64

PPMO # 10 acpP ATATCGCTCAC N ( RGR ) 4XB Acetyl 66

PPMO# 11 acpp ATATCGCTCAC N ( RYR ) 4XB - Acetyl 65

PPMO # 12 acpP ATATCGCTCAC N (RXR ) 4XB - H 59

PPMO # 13 acpP ?????cc???? W TEG ( RFF ) 3RXB 67 PPMO # 14 acpp CACAGGAATTC 4 TEG (RXR ) 4XB 59

PPMO # 15 acps TTGCCATTAGC U TEG ( RXR ) 4XB 59 PPMO # 16 acp - E CTGTAGTGATT TEG ( RXR ) 4XB 59 TCACCA O

PPMO # 17 fabA TTATCTACCAT 7 TEG ( RXR ) 4XB J PPMO # 18 fabB GCACGTTTCAT 0 TEG ( RXR ) 4XB 59

PPMO # 19 fabI AGAAAACCCAT 0 TEG ( RXR ) 4XB 59 PPMO # 20 gapa TTGATAGTCAT 10 TEG ( RXR ) 4XB 59 US 10 , 391 ,098 B2 57 58 TABLE 3A - continued Exemplary Fatty Acid Biosynthesis - Associated Targeting Sequences AONS

Targeting 5 ! 3 ! PMO Target Sequence TS SEQ Attachment Attachment CPP SEQ Name Gene ( TS ) ID NO : * * * * * ID NO . PPMO # 21 accA CTTTTTTCAT 11 (RXR ) 4XB - Acetyl 59 PPMO # 68 Scramble TCT CAG ATG 71 TEG ( RXR ) 2XB - 59 GT

The thymines ( T ) can be uracils ( U ) , and vice versa ; I is inosine ; * * X is 6 - aminohexanoic acid , B is beta - alanine , G is glycine , F is phenylalanine , Y is tyrosine , and TEG is defined above . * * * X is 6 - aminohexanoic acid , Bis beta - alanine , and a 5 ' CPP is linked through a pip - PDA moiety described above .

TABLE 3B Exemplary AONS targeting other biochemical pathways , cellular processes , and / or antibiotic resistance Targeting 51 3 ! PMO Target Sequence TS SEO Attachment Attachment CPP SEQ Name Gene ( TS ) * ID NO : * * * * * ID NO . PPMO # 22 murA ATCCATTTAGT 12 TEG ( RXR ) 4XB 59 PPMO # 23 mura CATTTAGTTTG 13 TEG ( RXR ) 4XB PPMO # 24 murA AATTTATCCAT 14 TEG ( RXR ) 4XB PPMO # 25 murA AAATTTATCCA 15 TEG ( RXR ) 4XB 59 PPMO # 26 rpmB ACTCGGGACAT 16 TEG ( RXR ) 4XB PPMO # 27 rpmB ??????????? TEG ( RXR ) 4XB PPMO # 28 rpmB GGCAGACTCGG 18 TEG (RXR ) 4XB PPMO # 29 rpmB CTTAGACATGG 19 TEG ( RXR ) 4XB PPMO # 30 adk ATGATACGCAT 20 TEG ( RXR ) XB 59 PPMO # 31 infA TCTTTGGCCAT 21 TEG ( RXR ) 4XB PPMO # 32 ftsz TCAAATGAGGC 22 ( RXR ) 4XB Acetyl PPMO # 33 ftsz TCAAATGAGGC 2 TEG ( RXR ) 4XB PPMO # 34 ftsz AATGAGGCCAT 3 TEG ( RXR ) 4XB PPMO # 35 ftsz ATAGTTTCTCT 24 (RXR ) 4XB Acetyl 59 CC PPMO # 36 rpoD TCATCTTTGCT 25 TEG ( RXR ) 4XB PPMO # 37 rpoD TTTTGCTCCAT 26 TEG ( RXR ) 4XB PPMO # 38 aroc ??cccTGCCAT 27 TEG ( RXR ) 4XB PPMO # 39 aroc TTTCCAGCCAT 28 TEG ( RXR ) 4XB PPMO # 40 murF ACGCTAATCAT 29 TEG ( RXR ) 4XB PPMO # 41 lpxC TGTTTGATCAT 30 TEG ( RXR ) 4XB PPMO # 42 kata AATTCGAGCAT TEG ( RXR ) 4XB PPMO # 43 boxA TGTTAAAGAGC 32 TEG (RXR ) 4XB PPMO # 44 rpoD - E CTTGTAACCAC 33 TEG (RXR ) 4XB ACCA PPMO # 45 pryc GGTGCAGTCAT 34 TEG ( RXR ) 4XB 59 PPMO # 46 pryA GACTTAATCAA 35 TEG ( RXR ) 4XB US 10 ,391 ,098 B2 59 60 TABLE 3B - continued Exemplary ANS targeting other biochemical pathways , cellular processes , and / or antibiotic resistance Targeting 3 ! PMO Target Sequence TS SEO Attachment Attachment CPP SEQ Name Gene ( TS ) * ID NO : * * * * * ID NO .

PPMO # 47 lgt CTACTGGTCAT 36 TEG ( RXR ) 4XB 59 PPMO # 48 folA CATTGAGATTT 37 TEG ( RXR ) XB PPMO # 49 infB ACATCTGTCAT 38 TEG ( RXR ) 4XB PPMO # 50 nrda TTCTGATTCAT 39 TEG ( RXR ) 4XB PPMO # 51 nrdB GTATATGCCAT 40 TEG ( RXR ) 4XB PPMO # 52 zipA TCCTGCATCAT TEG (RXR ) 4XB PPMO # 53 coaA ??????????? 4 TEG ( RXR ) 4XB PPMO # 54 qyrA - E GTTACCCTGAC 43 TEG ( RXR ) 4XB CGACCA PPMO # 55 gyrA - E GTTACCCTGAC 44 TEG ( RXR ) 4XB - 59 CACCA PPMO # 56 mrda TGTTTCATACG 45 TEG ( RXR ) 4XB 59 PPMO # 57 lpxB GGTTTGCCAAG 46 TEG (RXR ) 4XB PPMO # 58 lpxc TGTTTCACCAT 47 TEG ( RXR ) 4XB PPMO # 59 kdtA TTTTTCGCCAA 48 TEG ( RXR ) 4XB PPMO # 60 boxA CTCTTAATGAT 49 TEG ( RXR ) 4XB PPMO # 61 boxc ATCCACACAAG 50 TEG ( RXR ) 4XB PPMO # 62 rpoD - E TCCACCAAGTC 51 TEG ( RXR ) 4XB ACCA PPMO # 63 pryc AGAGTTCAAGG 52 TEG (RXR ) 4XB PPMO # 64 cara GGTGCTCAAAC 53 TEG ( RXR ) 4XB 59 PPMO # 65 adeA ATACTGTCCAA 54 TEG ( RXR ) 4XB PPMO # 66 blat ??????????? 55 TEG ( RXR ) 4XB PPMO # 67 cml TCCTTCTGATT 56 TEG (RXR ) 4XB 59 The thymines ( T ) can be uracils ( U ) , and vice versa ; I is inosine ; * * X is 6 - aminohexanoic acid , B is beta - alanine , Gis glycine , F is phenylalanine , Y is tyrosine , and TEG is defined above . * * * X is 6 - aminohexanoic acid , B is beta - alanine , and a 5 ' CPP is linked through a pip - PDA moiety described above . 50 IV . Methods of Use and Formulations compositions can further comprise one or more antimicro bial agents . The methods provided herein can be practiced in Embodiments of the present disclosure include methods vitro or in vivo . of using the antisense oligomers described herein to reduce For example , certain embodiments include methods of the expression and activity of one or more bacterial proteins treating a bacterial infection in a subject, comprising admin associated with biochemical pathways, cellular processes , 55 istering to a subject in need thereof ( e . g ., subject having or and /or antibiotic resistance . Certain embodiments include at risk for having a bacterial infection ) an antisense oligomer methods of using the antisense oligomers to reduce replica or pharmaceutical composition described herein . Also tion , proliferation , or growth of a bacteria , for example , to included are methods of reducing replication of a bacteria , treat a bacterial infection in a subject, either alone or in comprising contacting the bacterium with an antisense oli combination with one or more additional antimicrobial " gomer described herein . In some embodiments , the bacterium is selected from the agents . In some instances, the antisense oligomers increase genus Escherichia and Acinetobacter the susceptibility of the bacterium to one or more antimi Escherichia is a genus of Gram -negative , non -spore form crobial agents . ing, facultatively anaerobic , rod -shaped bacteria from the Also included are pharmaceutical compositions compris - 65 family Enterobacteriaceae , and includes the species Escheri ing the antisense oligomers , typically in combination with a chia coli , which is responsible for the vast majority of pharmaceutically -acceptable carrier . Certain pharmaceutical Escherichia -related pathogenesis . US 10 , 391, 098 B2 62 Acinetobacter is a genus of Gram -negative bacteria Examples of genes associated with biochemical pathways belonging to the class of Gammaproteobacteria . Examples and /or cellular processes include cellular homeostasis genes of clinically - relevant Acinetobacter complexes include the (and their related proteins) , for example, from Escherichia Acinetobacter calcoaceticus- baumannii complex (glucose species . In particular embodiments , the bacterium comprises oxidizing non hemolytic ) , Acinetobacter lwoffii ( glucose - 5 or expresses the adk gene , which encodes an adenylate negative non hemolytic ), and Acinetobacter haemolyticus kinase. In specific embodiments , the bacterium that com ( hemolytic ) . Specific examples include Acinetobacter bau - prises or expresses one or more genes associated with cellular homeostasis genes is Escherichia coli. mannii . Examples of genes associated with biochemical pathways Thus, in some embodiments , the bacterium is any of the 10 and /or cellular processes include protein biosynthesis genes foregoing members of the genera Escherichia or Acineto (and their related proteins ) , for example, from Escherichia bacter . In specific embodiments , the bacterium is Escheri species. In particular embodiments , the bacterium comprises chia coli or Acinetobacter baumannii . In some embodi or expresses the infA gene , which encodes a translation ments , the bacterium is selected from one or more of the initiation factor. In specific embodiments, the bacterium that strains in Table El. 15 comprises or expresses one or more genes associated with In certain embodiments , the bacterium is a multi- - drug proteinpro biosynthesis genes is Escherichia coli. resistance (MDR ) strain of bacteria . Multiple drug resistance Examples of genes associated with biochemical pathways (MDR ), multidrug resistance or multiresistance is a condi and / or cellular processes include cell division genes ( and tion enabling disease - causing microorganisms (bacteria , their related proteins ) , for example , from Acinetobacter spp . viruses, fungi or parasites) to resist distinct antimicrobials 20 In particular embodiments , the bacterium comprises or such as antibiotics, antifungal drugs, antiviral medications, expresses the ftsZ gene, which encodes a protein that antiparasitic drugs , and others . In particular embodiments , assembles into a ring at the future site of the septum of the bacterium is extensively - drug resistant (XDR ) or pan - bacterial cell division . In specific embodiments , the bacte drug resistant ( PDR ) . In some embodiments , the bacterium rium that comprises or expresses one or more genes asso is an extended - spectrum ß - lactamase (ESBLs ) producing 25 ciated with cell division genes is Acinetobacter spp . Gram - negative bacteria , or a multi - drug -resistant gram Examples of genes associated with biochemical pathways negative rod (MDR GNR ) MDRGN bacteria . In specific and /or cellular processes include RNA synthesis genes ( and embodiments , the bacterium is MDR Escherichia , for their related proteins ), for example , from Acinetobacter spp . example , MDR Escherichia coli , or MDR Acinetobacter , for In particular embodiments , the bacterium comprises or example , MDR Acinetobacter baumannii . 30 expresses the rpoD gene, which encodes a sigma D factor of Examples of genes associated with biochemical pathways RNA polymerase . In specific embodiments, the bacterium and / or cellular processes include fatty acid biosynthesis that comprises or expresses one or more genes associated genes ( and their related proteins ) such as acpp , acc? , acpS , with RNA synthesis genes is Acinetobacter spp . and / or fab genes , for example , from Escherichia or Acine Examples of genes associated with biochemical pathways tobacter . In particular embodiments , the bacterium com - 35 and / or cellular processes include aromatic compound bio prises or expresses the acpP gene , which encodes an acyl synthesis genes (and their related proteins ), for example , carrier protein . In particular embodiments , the bacterium from Acinetobacter spp . In particular embodiments , the comprises or expresses the accA gene , which encodes a bacterium comprises or expresses the aroC gene , which carboxyltransferase alpha subunit of an acetyl Coenzyme A encodes a chorismate synthase (5 - enolpyruvylshikimate - 3 carboxylase. In specific embodiments , the bacterium that 40 phosphate phospholyase ). In specific embodiments , the bac comprises or expresses one or more genes associated with terium that comprises or expresses one or more genes fatty acid biosynthesis is a Escherichia species, for example , associated with aromatic compound biosynthesis genes is Escherichia coli. In specific embodiments , the bacterium Acinetobacter spp . that comprises or expresses one or more genes associated In some embodiments , the bacteria or bacterium com with fatty acid biosynthesis is an Acinetobacter species . In 45 prises ( e . g . , encodes ) one or more antibiotic resistance some of these and related embodiments , the subject in need genes . General examples of antibiotic resistance genes (and thereof is immunocompromised and has an underlying lung their related proteins ) include beta - lactamases , which can disease , such as cystic fibrosis (CF ) or chronic granuloma- enzymatically deactivate certain antimicrobial agents , and tous disease (CGD ) . genes /proteins which increase the permeability or active Examples of genes associated with biochemical pathways 50 efflux ( pumping out) of an antimicrobial agent. Particular and / or cellular processes include peptidoglycan biosynthesis examples of antibiotic resistance genes include TEM beta genes (and their related proteins ) , for example , from lactamase (blaT ) , chloramphenicol resistance gene cml and Escherichia species. In particular embodiments , the bacte - resistance - nodulation - cell division (RND ) - type multidrug rium comprises or expresses the murA gene, which encodes efflux pump subunit AdeA ( adeA ) . In specific embodiments , a UDP -N -acetylglucosamine 1 -carboxyvinyltransferase . In 55 the bacterium is Escherichia coli or Acinetobacter spp . , specific embodiments , the bacterium that comprises or which comprises or expresses at least one antibiotic resis expresses one or more genes associated with peptidoglycan tance gene selected from blat, cml and adeA . biosynthesis is Escherichia coli. In some embodiments , the antisense oligomer reduces Examples of genes associated with biochemical pathways expression of the gene ( s ) associated with biochemical path and / or cellular processes include ribosomal protein genes 60 ways, cellular processes , and/ or antibiotic resistance in the ( and their related proteins ), for example, from Escherichia bacteria or bacterium . For instance , in some embodiments , species or Acinetobacter spp . In particular embodiments , the the antisense oligomer reduces expression by about or at bacterium comprises or expresses the rpmB gene, which least about 5 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , 60 , 65 , encodes a 50S ribosomal protein L28 . In specific embodi 70 , 75 , 80 , 85 , 90 , 95, 100 , 150 , 200 , 250 , 300 , 350 , 400 , ments, the bacterium that comprises or expresses one or 65 450 , 500 , 600 , 700 , 800 , 900 , or 1000 % or more ( including more genes associated with ribosomal protein genes is all integers and ranges in between ), relative to a control ( e . g . , Escherichia coli or Acinetobacter spp . absence of the antisense oligomer , scrambled oligomer, prior US 10 , 391 , 098 B2 63 64 to contacting with the oligomer ) , or by about or at least about treat (prophylactically or therapeutically ) an infection by 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , any of the bacteria described herein . In conjunction with 60 , 65 , 70 , 75 , 80 , 85, 90 , 95 , 100 , 200 , 300 , 400 , 500 , such treatment, pharmacogenomics ( e . g ., the study of the 1000 - fold or more (including all integers and ranges in relationship between an individual' s genotype /phenotype between ) , relative to a control. In some embodiments , the 5 and that individual' s response to a foreign compound or antisense oligomer reduces expression of one or more of drug ) may be considered . Differences in metabolism of AcpP , ACCA , MurA , RpmB , Adk , InfA , FtsZ , RpoD , AroC , therapeutics can lead to severe toxicity or therapeutic failure BlaT, Cml and / or AdeA and the bacterium is an Acineto - by altering the relation between dose and blood concentra bacter or Escherichia species which comprises or expresses tion of the pharmacologically active drug . one or more of AcpP , ACCA , MurA , RpmB , Adk , InfA , FtsZ , 10 Thus , a physician or clinician may consider applying Rpod , AroC , Blat , Cml and / or AdeA . Gene or protein knowledge obtained in relevant pharmacogenomics studies expression can bemeasured in vitro ( see , e . g ., the Examples ) in determining whether to administer a therapeutic agent as or in vivo . well as tailoring the dosage and /or therapeutic regimen of In some embodiments, the antisense oligomer reduces or treatment with a therapeutic agent. inhibits the growth of the bacteria or bacterium . For 15 Effective delivery of the antisense oligomer to the target instance , in some embodiments , the antisense oligomer nucleic acid is one aspect of treatment. Routes of antisense reduces growth of the bacteria or bacterium by about or at oligomer delivery include , but are not limited to , various least about 5 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , 60 , 65, systemic routes, including oral and parenteral routes, e. g ., 70 , 75 , 80 , 85 , 90 , 95 , 100 , 150 , 200 , 250 , 300 , 350 , 400 , intravenous, subcutaneous, intraperitoneal , and intramuscu 450 , 500 , 600 , 700 , 800 , 900 , or 1000 % or more ( including 20 lar, as well as inhalation , transdermal, and topical delivery . all integers and ranges in between ) , relative to a control ( e . g ., The appropriate route may be determined by one of skill in absence of the antisense oligomer, scrambled oligomer, prior the art , as appropriate to the condition of the subject under to contacting with the oligomer ), or by about or at least about treatment. Vascular or extravascular circulation , the blood or 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , lymph system , and the cerebrospinal fluid are some non 60 , 65 , 70 , 75, 80 , 85 , 90 , 95 , 100 , 200 , 300 , 400 , 500 , 25 limiting sites where the antisense oligomers may be intro 1000 - fold or more ( including all integers and ranges in duced . Direct CNS delivery may be employed , for instance , between ), relative to a control. Bacterial growth can be intracerebral, intraventricular, or intrathecal administration measured in vitro (see , e . g . , the Examples ) or in vivo . In may be used as routes of administration . particular embodiments , the antisense oligomer that reduces In certain embodiments , the antisense oligomers can be growth of the bacterium is targeted against a protein asso - 30 delivered by transdermal methods ( e . g . , via incorporation of ciated with a biochemical pathway and / or cellular process the antisense oligomers into , e . g ., emulsions , with such selected from one or more of AcpP , ACCA , MurA , RpmB , antisense oligomers optionally packaged into liposomes ) . Adk , InfA , FtsZ , RpoD , AroC , BlaT, Cml and /or AdeA and Such transdermal and emulsion / liposome- mediated methods the bacterium is an Acinetobacter or Escherichia species of delivery are described for delivery of antisense oligomers which comprises or expresses one or more of AcpP, ACCA , 35 in the art, e . g . , in U . S . Pat. No. 6 , 965 ,025 , the contents of MurA , RpmB , Adk , InfA , FtsZ , RpoD , AroC , BlaT, Cml which are incorporated in their entirety by reference herein . and / or AdeA . In some embodiments , as described herein , the The antisense oligomers described herein may also be antisense oligomer is employed in combination with one or delivered via an implantable device. Design of such a device more antimicrobial agents, for example , to synergistically is an art - recognized process, with , e . g ., synthetic implant reduce the growth of the bacteria or bacterium . 40 design described in , e. g ., U . S . Pat . No . 6 , 969, 400 , the In some embodiments, the antisense oligomer reduces contents of which are incorporated by reference . beta -lactamase ( e . g ., carbapenemase ) activity in the peri - Antisense oligomers can be introduced into cells using plasm of the bacterium by about or at least about 5 , 10 , 15 , art - recognized techniques ( e . g . , transfection , electropora 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , 60 , 65 , 70 , 75 , 80 , 85 , 90 , 95 , tion , fusion , liposomes , colloidal polymeric particles and 100 , 150 , 200 , 250 , 300 , 350 , 400 , 450 , 500 , 600 , 700 , 800 , 45 viral and non -viral vectors as well as other means known in 900 , or 1000 % or more ( including all integers and ranges in the art ). The method of delivery selected will depend at least between ), relative to a control, or by at least about 2 , 3 , 4 , on the oligomer chemistry , the cells to be treated and the 5 , 6 , 7 , 8 , 9 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , 60 , 65 , location of the cells and will be apparent to the skilled 70 , 75 , 80 , 85 , 90 , 95 , or 100 - fold or more ( including all artisan . For instance , localization can be achieved by lipo integers and ranges in between ), relative to a control. In 50 somes with specific markers on the surface to direct the some embodiments, the antisense oligomer reduces mero - liposome, direct injection into tissue containing target cells , penemase enzymatic activity in the periplasm of the bacte - specific receptor -mediated uptake , or the like . rium . In particular embodiments , the antisense oligomer that As known in the art , antisense oligomers may be delivered reduces beta - lactamase activity is targeted against blat , and using , e. g ., methods involving liposome- mediated uptake , the bacterium is an Acinetobacter , or Escherichia species, 55 lipid conjugates , polylysine -mediated uptake, nanoparticle for example , Escherichia coli or Acinetobacter baumannii mediated uptake, and receptor- mediated endocytosis , as well which comprises or expresses BlaT . These are exemplary as additional non - endocytic modes of delivery , such as bacterial species and it is expected that any bacterium microinjection , permeabilization ( e . g . , streptolysin - o per expressing the blat gene is susceptible to the compounds meabilization , anionic peptide permeabilization ) , electropo and methods described herein . Beta -lactamase activity can 60 ration , and various non - invasive non -endocytic methods of be measured according to routine techniques in the art. delivery that are known in the art ( see , e . g . , Dokka and In some embodiments , the methods are practiced in vivo , Rojanasakul, Advanced Drug Delivery Reviews 44 : 35 -49 , and comprise administering the antisense oligomer to a incorporated by reference in its entirety ) . subject in need thereof, for example , a subject in need The antisense oligomers may be administered in any thereof that is infected or at risk for being infected by one or 65 convenient vehicle or carrier which is physiologically and /or more of the bacteria described herein . The antisense oligom - pharmaceutically acceptable . Such a composition may ers described herein can thus be administered to subjects to include any of a variety of standard pharmaceutically US 10 , 391 , 098 B2 65 66 acceptable carriers employed by those of ordinary skill in 93/ 01286 . Alternatively, the oligomers may be administered the art . Examples include , but are not limited to , saline , in microspheres or microparticles. (See , e . g . , Wu , G . Y . and phosphate buffered saline (PBS ) , water, aqueous ethanol, Wu, C . H ., J. Biol. Chem . 262 :4429 -4432 , 30 1987 ) . Alter emulsions, such as oil/ water emulsions or triglyceride emul natively , the use of gas- filled microbubbles complexed with sions , tablets and capsules . The choice of suitable physi - 5 the antisense oligomers can enhance delivery to target ologically acceptable carrier will vary dependent upon the tissues , as described in U . S . Pat . No . 6 , 245 , 747. Sustained chosen mode of administration . “ Pharmaceutically accept release compositions may also be used . These may include able carrier” is intended to include any and all solvents , dispersion media , coatings , antibacterial and antifungal semipermeable polymeric matrices in the form of shaped agents , isotonic and absorption delaying agents , and the like , 10 articles such as films or microcapsules. compatible with pharmaceutical administration . The use of In certain embodiments , the antisense oligomer is admin such media and agents for pharmaceutically active sub istered to a mammalian subject , e . g ., human or domestic stances is well known in the art. Except insofar as any animal, exhibiting the symptoms of a bacterial infection conventionalmedia or agent is incompatible with the active ( e . g ., antibiotic resistance or MDR bacterial infection ) , in a compound , use thereof in the compositions is contemplated . 15 suitable pharmaceutical carrier . In some aspects , the subject Supplementary active compounds can also be incorporated is a human subject, e . g ., a patient diagnosed as having a into the compositions bacterial infection . In particular embodiments , the antisense The compounds ( e . g ., antisense oligomers , antimicrobial oligomer is contained in a pharmaceutically acceptable agents ) described herein may generally be utilized as the carrier , and is delivered orally . In some embodiments , the free acid or free base . Alternatively , the compounds 20 antisense oligomer is contained in a pharmaceutically described herein may be used in the form of acid or base acceptable carrier, and is delivered intravenously ( i . v . ) . addition salts . Acid addition salts of the free amino com In some embodiments , the antisense oligomer is admin pounds described herein may be prepared by methods well istered in an amount and manner effective to result in a peak known in the art , and may be formed from organic and blood concentration of at least 200 -400 nM antisense oli inorganic acids. Suitable organic acids include maleic, 25 gomer. Typically , one or more doses of antisense oligomer fumaric, benzoic , ascorbic , succinic , methanesulfonic, ace - are administered , generally at regular intervals , for a period tic , trifluoroacetic , oxalic , propionic , tartaric , salicylic , cit of about one to two weeks . Certain doses for oral adminis ric , gluconic , lactic , mandelic , cinnamic , aspartic , stearic , tration are from about 1 - 1000 mg oligomer per 70 kg . In palmitic, glycolic , glutamic , and benzenesulfonic acids. some cases , doses of greater than 1000 mg oligomer /patient Suitable inorganic acids include hydrochloric , hydrobro - 30 mic , sulfuric , phosphoric , and nitric acids. Base addition may be necessary . For i . v . administration , some doses are salts included those salts that form with the carboxylate from about 0 . 5 mg to 1000 mg oligomer per 70 kg. The anion and include salts formed with organic and inorganic antisense oligomer may be administered at regular intervals cations such as those chosen from the alkali and alkaline for a short time period , e .g ., daily for two weeks or less . earth metals ( for example , lithium , sodium , potassium , mag - 35 However, in some cases the antisense oligomer is adminis nesium , barium and calcium ) , as well as the ammonium ion tered intermittently over a longer period of time. Adminis and substituted derivatives thereof ( for example , dibenzy tration may be followed by, or concurrent with , administra lammonium , benzylammonium , 2 -hydroxyethylammonium , tion of an antimicrobial ( e . g ., antibiotic ) or other therapeutic and the like ). Thus , the term “ pharmaceutically acceptable treatment, as described herein . The treatment regimen may salt ” is intended to encompass any and all acceptable salt 40 be adjusted ( dose , frequency , route , etc . ) as indicated , based forms. on the results of immunoassays , other biochemical tests and In addition , prodrugs are also included within the context physiological examination of the subject under treatment. of this disclosure . Prodrugs are any covalently bonded An effective in vivo treatment regimen using the antisense carriers that release a compound in vivo when such prodrug o ligomers described herein may vary according to the dura is administered to a patient. Prodrugs are generally prepared 45 tion , dose , frequency and route of administration , as well as by modifying functional groups in a way such that the the condition of the subject under treatment (i . e ., prophy modification is cleaved , either by routine manipulation or in lactic administration versus administration in response to vivo , yielding the parent compound . Prodrugs include , for localized or systemic infection ) . Accordingly , such in vivo example , compounds of this disclosure wherein hydroxy , therapy will often include monitoring by tests appropriate to amine or sulfhydryl groups are bonded to any group that , 50 the particular type of disorder or bacterial infection under when administered to a patient , cleaves to form the hydroxy, treatment, and corresponding adjustments in the dose or amine or sulfhydryl groups. Thus , representative examples treatment regimen , in order to achieve an optimal therapeu of prodrugs include (but are not limited to ) acetate , formate tic outcome. and benzoate derivatives of alcohol and amine functional Treatment may be monitored , e . g . , by general indicators groups of the antisense oligomers described herein . Further , 55 of disease known in the art. The efficacy of an in vivo in the case of a carboxylic acid ( COOH ) , esters may be administered antisense oligomer described herein may be employed , such as methyl esters , ethyl esters , and the like . determined from biological samples (tissue , blood , urine In some instances , liposomes may be employed to facili - etc . ) taken from a subject prior to , during and subsequent to tate uptake of the antisense oligomer into cells ( see , e . g . , administration of the antisense oligomer . Assays of such Williams, S . A ., Leukemia 10 ( 12 ): 1980 - 1989 , 1996 ; Lap - 60 samples include ( 1) monitoring the presence or absence of palainen et al. , Antiviral Res. 23 : 119 , 1994 ; Uhlmann et al. , heteroduplex formation with target and non - target antisense oligomers : a new therapeutic principle , Chemical sequences , using procedures known to those skilled in the Reviews, Volume 90 , No . 4 , 25 pages 544 -584 , 1990 ; art, e . g . , an electrophoretic gel mobility assay ; ( 2 ) monitor Gregoriadis , G ., Chapter 14 , Liposomes , Drug Carriers in ing the amount of a mutant mRNA in relation to a reference Biology and Medicine, pp . 287 - 341, Academic Press, 1979 ) . 65 normal mRNA or protein as determined by standard tech Hydrogels may also be used as vehicles for antisense niques such as RT- PCR , Northern blotting , ELISA or West oligomer administration , for example , as described in WO ern blotting . US 10 , 391 , 098 B2 67 68 V . Combination Therapies pyridazine , sulfametoxydiazine , sulfadoxine, and Certain embodiments include combination therapies , for sulfametopyrazine ; quinolones such as cinoxacin , nalidixic example, the administration of antisense oligomers in com acid , oxolinic acid (Uroxin ) , piromidic acid (Panacid ) , bination with antimicrobial agents such as antibiotics. Com pipemidic acid (Dolcol ) rosoxacin ( Eradacil ), ciprofloxacin bination therapies can be employed , for example , to increase 5 ( Alcipro , Ciprobay, Cipro , Ciproxin , ultracipro ), enoxacin the sensitivity or susceptibility of a given bacteria to one or ( Enroxil, Penetrex ) , fleroxacin (Megalone , Roquinol) , lom more antimicrobial agents , and thereby improve the thera efloxacin (Maxaquin ) , nadifloxacin ( Acuatim , Nadoxin , peutic outcome ( e . g . , resolution of the infection ) . Likewise , certain combination therapies can be employed , for Nadixa ) , norfloxacin (Lexinor , Noroxin , Quinabic , Janacin ), example , to reduce or reverse the resistance of a given 10 ofloxacin (Floxin , Oxaldin , Tarivid ), pefloxacin (Peflacine ) , bacteria to one or more antimicrobial agents . In particular rufloxacin (Uroflox ) , balofloxacin (Baloxin ) , grepafloxacin embodiments , the antisense oligomer reduces the minimum ( Raxar ) , levofloxacin ( Cravit, Levaquin , Tavanic ) , pazu inhibitory concentration (MIC ) of an antibiotic against a floxacin ( Pasil, Pazucross ) , sparfloxacin (Zagam ), tema given bacterium . In certain embodiments , the antisense floxacin (Omniflox ), tosufloxacin (Ozex , Tosacin ) , clina oligomer and the antimicrobial agent display synergy in 15 floxacinTox , gatifloxacin ( Zigat, Tequin (Zymar - opth . ) , reducing bacterial growth and / or increasing bacterial cell gemifloxacin (Factive ) , moxifloxacin ( Acflox Woodward , killing . Also included are pharmaceutical compositions, as Avelox , Vigamox , sitafloxacin (Gracevit ) , trovafloxacin described herein , which comprise an antisense oligomer and ( Trovan ), prulifloxacin ( Quisnon ); oxazolidinones such as an antimicrobial agent such as antibiotic . eperezolid , linezolid , posizolid , radezolid , ranbezolid , sut In some embodiments , the antisense oligomer and the 20 ezolid , and tedizolid ; polymyxins such as polysporin , neo antimicrobial agent are administered separately . In certain sporin , polymyxin B , polymyxin E ( colistin ) ; rifamycins embodiments , the antisense oligomer and the antimicrobial such as rifampicin or rifampin , rifabutin , rifapentine , and agent are administered sequentially . In some embodiments , rifaximin ; lipiarmycins such as fidaxomicin ; macrolides the antisense oligomer and the antimicrobial agent are such as azithromycin , clarithromycin , dirithromycin , eryth administered concurrently , for example , as part of the same 25 romycin , roxithromycin , telithromycin , carbomycin A , or different pharmaceutical composition . josamycin , kitasamycin , midecamycin /midecamycin Examples of antimicrobial agents ( e . g ., antibiotics ) that acetate, oleandomycin , solithromycin , spiramycin , and tro can be administered in combination with an antisense oli - leandomycin ; lincosamides such as lincomycin , clindamy gomer include beta - lactam antibiotics such as carbapenems, cin , and pirlimycin ; cyclic lipopeptides such as daptomycin ; penicillin and penicillin derivatives ( or penams) , ampicillin , 30 glycopeptides such as vancomycin and teichoplanin ; glycyl chloramphenicol, cephalosporins ( e . g . , Cefacetrile cyclines such as tigecycline . Thus, any one or more of the ( cephacetrile ), Cefadroxil ( cefadroxyl; Duricef) , Cephalexin foregoing antibiotics can be combined with any of the ( cefalexin ; Keflex ) , Cefaloglycin ( cephaloglycin ), Cefalo - antisense oligomers described herein , for the treatment of nium (cephalonium ), Cefaloridine (cephaloridine ), Cefalo - any of the bacterium or bacteria described herein . tin ( cephalothin ; Keflin ), Cefapirin (cephapirin ; Cefadryl) , 35 In some embodiments , the antimicrobial agent is selected Cefatrizine , Cefazaflur, Cefazedone , Cefazolin (cephazolin ; from one or more of aminoglycoside antibiotics, tetracycline Ancef, Kefzol) , Cefradine ( cephradine; Velosef ), Cefroxa antibiotics, and ß - lactam antibiotics, as described herein . In dine , Ceftezole , Cefaclor ( Ceclor, Distaclor, Keflor, Rani some of these and related embodiments , the bacterium clor ) , Cefonicid (Monocid ) , Cefprozil (cefproxil ; Cefzil ) , comprises or expresses a gene selected from one or more of Cefuroxime ( Zefu , Zinnat, Zinacef, Ceftin , Biofuroksym , 40 accA , and the antisense oligomer is targeted against the fatty Xorimax ) , Cefuzonam , Cefmetazole , Cefotetan , Cefoxitin , acid biosynthesis gene . In some of these and related embodi loracarbef (Lorabid ); Cephamycins: cefbuperazone , cef ments , the bacterium comprises or expresses a gene selected metazole (Zefazone ) , cefminox , cefotetan (Cefotan ) , cefoxi- from one or more of murA , and the antisense oligomer is tin (Mefoxin ), Cefotiam (Pansporin ) , Cefcapene, Cefdalox - targeted against the peptidoglycan biosynthesis gene . In ime, Cefdinir (Sefdin , Zinir , Omnicef , Kefnir ) , Cefditoren , 45 some of these and related embodiments , the bacterium Cefetamet, Cefixime ( Fixx , Zifi , Suprax ), Cefmenoxime, comprises or expresses a gene selected from one or more of Cefodizime, Cefotaxime ( Claforan ) , Cefovecin ( Convenia ) , rpmB , and the antisense oligomer is targeted against the Cefpimizole , Cefpodoxime (Vantin , PECEF ) , Cefteram , ribosomal protein gene. In some of these and related Ceftibuten (Cedax ) , Ceftiofur, Ceftiolene , Ceftizoxime embodiments , the bacterium comprises or expresses a gene (Cefizox ), Ceftriaxone (Rocephin ), Cefoperazone (Cefobid ) , 50 selected from one or more of adk , and the antisense oligomer Ceftazidime (Meezat , Fortum , Fortaz ), latamoxef (moxa - is targeted against the cellular homeostasis gene . In some of lactam ) , Cefclidine, cefepime (Maxipime ) , cefluprenam , these and related embodiments , the bacterium comprises or cefoselis , Cefozopran , Cefpirome (Cefrom ) , Cefquinome, expresses a gene selected from one or more of infA , and the flomoxef, Ceftobiprole , Ceftaroline, Cefaloram , Cefaparole , antisense oligomer is targeted against the protein biosynthe Cefcanel, Cefedrolor, Cefempidone , Cefetrizole , Cefivitril , 55 sis gene. In some of these and related embodiments , the Cefmatilen , Cefmepidium , Cefoxazole , Cefrotil, Cefsumide , bacterium comprises or expresses a gene selected from one Ceftioxide , Cefuracetime) , and monobactams ( e . g . , aztreo - or more of ftsZ , and the antisense oligomer is targeted nam , tigemonam , nocardin A , tabtoxin ) ; aminoglycosides against the cell division gene . In some of these and related such as tobramycin , gentamicin , kanamycin a , amikacin , embodiments , the bacterium comprises or expresses a gene dibekacin , sisomicin , netilmicin , neomycin B , neomycin C , 60 selected from one or more of rpon , and the antisense neomycin E ( paromomycin ) , and streptomycin ; tetracy - oligomer is targeted against the RNA synthesis gene . In clines such as tetracycline , chlortetracycline, oxytetracy - some of these and related embodiments , the bacterium cline , demeclocycline, lymecycline , meclocycline , methacy comprises or expresses a gene selected from one or more of cline , minocycline , rolitetracycline , and doxycyline ; aroC , and the antisense oligomer is targeted against the sulfonamides such as sulfacetamide , sulfadiazine , sulfadimi- 65 aromatic compound biosynthesis gene. In specific embodi dine , sulfafurazole , sulfisomidine , sulfadoxine , sulfame- ments , the bacterium is Escherichia coli or Acinetobacter thoxazole , sulfamoxole , sulfadimethoxine , sulfamethoxy spp . US 10 , 391 , 098 B2 69 70 In some embodiments , the antimicrobial agent is a beta - bacteria or bacterium to the antimicrobial agent by increas lactam antibiotic , as described herein . In certain of these and ing the bactericidal (cell -killing ) and / or bacteriostatic related embodiments, the bacterium comprises or expresses ( growth - slowing ) activity of the antimicrobial agent against a beta - lactamase such as BlaT, and the antisense oligomer is the bacteria or bacterium being targeted , relative to the targeted against the beta - lactamase . In particular embodi- 5 antimicrobial agent alone . In particular embodiments , the ments , the antimicrobial agent is a carbapenem . Examples of antisense oligomer increases the susceptibility or sensitivity carbapenems include meropenem , imipenem , ertapenem , by about or at least about 5 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , doripenem , panipenem , biapenem , razupenem , tebipenem , 50 , 55 , 60 , 65 , 70 , 75 , 80 , 85 , 90 , 95 , 100 , 150 , 200 , 250 , lenapenem , tomopenem , and ampicillin . In specific embodi - 300 , 350 , 400 , 450 , 500 , 600 , 700 , 800 , 900 , or 1000 % or ments, the antimicrobial agent is meropenem . In particular 10 more ( including all integers and ranges in between ) , relative embodiments , the antimicrobial agent is a cephalosporin to the antimicrobial agent alone , or by about or at least about ( cephem ), penicillin or penicillin derivative (penam ). In 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 15 , 20, 25 , 30 , 35 , 40 , 45 , 50 , 55 , particular embodiments , the antisense oligomer reduces the 60 , 65 , 70 , 75 , 80 , 85 , 90 , 95 , 100 , 200 , 300 , 400 , 500 , MIC of a carbapenem such asmeropenem against a bacteria , 1000 - fold or more ( including all integers and ranges in for example , a MDR strain of E . coli or Acinetobacter 15 between ), relative to the antimicrobial agent alone . In some baumannii. In some embodiments , the combination of the embodiments , the antisense oligomer synergistically antisense oligomer and the carbapenem such as meropenem increases the susceptibility or sensitivity of a given bacteria reduces ( e . g . , synergistically reduces ) bacterial cell growth to the antimicrobial agent, relative to the antimicrobial agent or increase ( e . g . , synergistically increases ) bacterial cell - alone . killing , for example , of a MDR strain of E . coli or Acine - 20 In some embodiments , the antisense oligomer reduces the tobacter baumannii . minimum inhibitory concentration (MIC ) of an antimicro In some embodiments , the antimicrobial agent is an bial agent against the bacteria or bacterium being targeted , aminoglycoside , as described herein . Examples of amino - relative to the antimicrobial agent alone . The “ minimum glycosides include tobramycin , gentamicin , kanamycin a , inhibitory concentration ” or “ MIC ” refers to the lowest amikacin , dibekacin , sisomicin , netilmicin , neomycin B , 25 concentration of an antimicrobial agent that will inhibit the neomycin C , neomycin E ( paromomycin ), and streptomycin . visible growth of a microorganism after overnight ( in vitro ) In specific embodiments , the antimicrobial agent is tobramy incubation . Minimum inhibitory concentrations are impor cin . In particular embodiments , the antisense oligomer tant in diagnostic laboratories to confirm resistance of reduces the MIC of an aminoglycoside such as tobramycin microorganismsto an antimicrobial agent and also to moni against a bacteria , for example , a MDR strain of E . coli or 30 tor the activity of new antimicrobial agents . The MIC is Acinetobacter baumannii . In some embodiments, the com - generally regarded as the most basic laboratory measure bination of the antisense oligomer and the aminoglycoside ment of the activity of an antimicrobial agent against a such as tobramycin reduces ( e . g . , synergistically reduces ) bacterial organism . Thus , in certain embodiments , the oli bacterial cell growth or increases ( e . g . , synergistically gomer reduces the minimum inhibitory concentration (MIC ) increases ) bacterial cell- killing , for example , of a MDR 35 of an antimicrobial agent against the bacteria or bacterium strain of E . coli or Acinetobacter baumannii . In some of by at least about 5 , 10 , 15 , 20 , 25 , 30 , 35 , 40 , 45 , 50 , 55 , 60 , these and related embodiments , the bacterium comprises or 65 , 70 , 75 , 80 , 85 , 90 , 95 , 100 , 150 , 200 , 250 , 300 , 350 , 400 , expresses the antibiotic resistance gene ade A , and the anti - 450 , 500 , 600 , 700 , 800 , 900 , or 1000 % or more ( including sense oligomer is targeted against the antibiotic resistance all integers and ranges in between ) , relative to the antimi gene . In specific embodiments, the bacterium is Escherichia 40 crobial agent alone . In certain embodiments , the oligomer coli or Acinetobacter baumannii . reduces the minimum inhibitory concentration (MIC ) of an In certain embodiments , the antimicrobial agent is a antimicrobial agent against the bacteria or bacterium by polymyxin such as colistin (polymyxin E ) , polysporin , neo - about or at least about 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 15 , 20 , 25 , sporin , or polymyxin B . In specific embodiments, the anti - 30 , 35 , 40 , 45 , 50 , 55 , 60 , 65 , 70 , 75 , 80 , 85 , 90 , 95 , 100 , 200 , microbial agent is colistin . In particular embodiments , the 45 300 , 400, 500 , 1000 - fold or more ( including all integers and antisense oligomer reduces the MIC of a polymyxin such as ranges in between ), relative to the antimicrobial agent alone . colistin against a bacteria , for example , a MDR strain of E . In some embodiments , the antisense oligomer synergisti coli or Acinetobacter baumannii. In some embodiments , the cally reduces the MIC of an antimicrobial agent against the combination of the antisense oligomer and the polymyxin bacteria or bacterium being targeted , relative to the antimi such as colistin reduces ( e . g . , synergistically reduces ) bac - 50 crobial agent alone . terial cell growth or increases ( e . g ., synergistically In some embodiments, the antisense oligomer that increases ) bacterial cell- killing , for example , of a MDR increases the sensitivity or reduces the MIC is targeted strain of E . coli or Acinetobacter baumannii . against blat, the bacterium is Escherichia coli or Acineto In certain embodiments, the antimicrobial agent includes bacter spp . that comprises or expresses Blat , and the one or more of ceftazidime, doxycycline , piperacillin ,mero - 55 antimicrobial agent is a beta - lactam such as cephalosporins , penem , chloramphenicol , and / or co - trimoxazole (trimethop - penicillin , penicillin derivatives ( penams) or ampicillin . rim /sulfamethoxazole ). In some of these and related In some embodiments, the antisense oligomer that embodiments , the bacterium is an Escherichia species that increases the sensitivity or reduces the MIC is targeted comprises or expresses one or more antibiotic resistance against cml, the bacterium is Escherichia coli or Acineto genes such as cml, and the antisense oligomer is targeted 60 bacter spp . that comprises or expresses Cml, and the anti against the antibiotic resistance gene( s ) . In specific embodi- microbial agent is chloramphenicol. ments , the bacterium is Escherichia coli . In particular embodiments, the antisense oligomer that In some embodiments, the antisense oligomer increases increases the sensitivity or reduces the MIC is targeted the susceptibility or sensitivity of a given bacterium to the against adeA , the bacterium is Escherichia coli or Acineto antimicrobial agent, relative to the antimicrobial agent 65 bacter baumannii that comprises or expresses AdeA , and the alone . For example , in certain embodiments , the antisense antimicrobial agent is an aminoglycoside antibiotic ( e . g . , oligomer increases the susceptibility or sensitivity of the tobramycin , gentamicin , kanamycin , amikacin , dibekacin , US 10 , 391, 098 B2 71 72 sisomicin , netilmicin , neomycin B , neomycin C , neomycin ( e . g ., tetracycline , chlortetracycline , oxytetracycline , deme E (paromomycin ), streptomycin ), a tetracycline antibiotic clocycline, lymecycline , meclocycline , methacycline , mino ( e . g . , tetracycline , chlortetracycline , oxytetracycline , deme - cycline , rolitetracycline , doxycyline ) , or a B - lactam antibi clocycline , lymecycline, meclocycline , methacycline , mino otic ( e . g ., carbapenem , penicillin derivative ( penam ) , cycline , rolitetracycline , doxycyline ) , or a B - lactam antibi- 5 cephalosporin ( cephem ) , monobactam ) . otic ( e . g . , carbapenem , penicillin derivative ( penam ) , In particular embodiments , the antisense oligomer that cephalosporin ( cephem ) , monobactam ) . increases the sensitivity or reduces the MIC is targeted In particular embodiments , the antisense oligomer that against fts Z , the bacterium is Escherichia coli or Acineto increases the sensitivity or reduces the MIC is targeted bacter baumannii that comprises or expresses FtsZ , and the against acc? , the bacterium is Escherichia coli or Acineto - 10 antimicrobial agent is an aminoglycoside antibiotic ( e . g . , bacter baumannii that comprises or expresses AccA , and the tobramycin , gentamicin , kanamycin , amikacin , dibekacin , antimicrobial agent is an aminoglycoside antibiotic ( e . g . , sisomicin , netilmicin , neomycin B , neomycin C , neomycin tobramycin , gentamicin , kanamycin , amikacin , dibekacin , E (paromomycin ) , streptomycin ), a tetracycline antibiotic sisomicin , netilmicin , neomycin B , neomycin C , neomycin (e .g ., tetracycline , chlortetracycline, oxytetracycline , deme E (paromomycin ) , streptomycin ) , a tetracycline antibiotic 15 clocycline, lymecycline , meclocycline, methacycline , mino ( e . g ., tetracycline , chlortetracycline , oxytetracycline, deme cycline, rolitetracycline, doxycyline ), or a B -lactam antibi clocycline , lymecycline , meclocycline, methacycline , mino - otic (e .g ., carbapenem , penicillin derivative (penam ) , cycline , rolitetracycline, doxycyline ) , or a ß - lactam antibi- cephalosporin ( cephem ) , monobactam ). otic ( e . g . , carbapenem , penicillin derivative ( penam ), In particular embodiments , the antisense oligomer that cephalosporin (cephem ), monobactam ) . 20 increases the sensitivity or reduces the MIC is targeted In particular embodiments , the antisense oligomer that against rpoD , the bacterium is Escherichia coli or Acineto increases the sensitivity or reduces the MIC is targeted bacter baumannii that comprises or expresses Rpod , and the against murA , the bacterium is Escherichia coli or Acine - antimicrobial agent is an aminoglycoside antibiotic ( e. g ., tobacter baumannii that comprises or expresses MurA , and tobramycin , gentamicin , kanamycin , amikacin , dibekacin , the antimicrobial agent is an aminoglycoside antibiotic ( e . g ., 25 sisomicin , netilmicin , neomycin B , neomycin C , neomycin tobramycin , gentamicin , kanamycin , amikacin , dibekacin , E (paromomycin ), streptomycin ), a tetracycline antibiotic sisomicin , netilmicin , neomycin B , neomycin C , neomycin (e . g ., tetracycline, chlortetracycline , oxytetracycline , deme E (paromomycin ) , streptomycin ) , a tetracycline antibiotic clocycline, lymecycline , meclocycline, methacycline , mino ( e . g . , tetracycline , chlortetracycline, oxytetracycline , deme - cycline, rolitetracycline , doxycyline ), or a B - lactam antibi clocycline , lymecycline ,meclocycline , methacycline, mino - 30 otic ( e . g ., carbapenem , penicillin derivative (penam ) , cycline, rolitetracycline, doxycyline ), or a B -lactam antibi otic ( e . g . , carbapenem , penicillin derivative ( penam ), In particular embodiments , the antisense oligomer that cephalosporin (cephem ) , monobactam ). increases the sensitivity or reduces the MIC is targeted In particular embodiments , the antisense oligomer that against aroC , the bacterium is Escherichia coli or Acineto increases the sensitivity or reduces the MIC is targeted 35 bacter baumannii that comprises or expresses AroC , and the against rpmB, the bacterium is Escherichia coli or Acine - antimicrobial agent is an aminoglycoside antibiotic ( e . g . , tobacter baumannii that comprises or expresses RpmB , and tobramycin , gentamicin , kanamycin , amikacin , dibekacin , the antimicrobial agent is an aminoglycoside antibiotic ( e . g . , sisomicin , netilmicin , neomycin B , neomycin C , neomycin tobramycin , gentamicin , kanamycin , amikacin , dibekacin , E (paromomycin ) , streptomycin ) , a tetracycline antibiotic sisomicin , netilmicin , neomycin B , neomycin C , neomycin 40 ( e . g ., tetracycline, chlortetracycline , oxytetracycline , deme E (paromomycin ) , streptomycin ) , a tetracycline antibiotic clocycline , lymecycline , meclocycline, methacycline, mino ( e . g . , tetracycline, chlortetracycline , oxytetracycline, deme- cycline , rolitetracycline , doxycyline ), or a B - lactam antibi clocycline , lymecycline , meclocycline , methacycline , mino - otic ( e . g . , carbapenem , penicillin derivative (penam ) , cycline, rolitetracycline , doxycyline ) , or a ß - lactam antibi- cephalosporin (cephem ), monobactam ) . otic ( e . g . , carbapenem , penicillin derivative ( penam ), 45 VI. Treatment Monitoring Methods cephalosporin ( cephem ), monobactam ). The efficacy of a given therapeutic regimen involving the In particular embodiments, the antisense oligomer that methods described herein may be monitored , for example , increases the sensitivity or reduces the MIC is targeted by general indicators ofbacterial infection , such as complete against adk , the bacterium is Escherichia coli or Acineto - blood count (CBC ) , nucleic acid detection methods, immu bacter baumannii that comprises or expresses Adk , and the 50 nodiagnostic tests , or bacterial culture . antimicrobial agent is an aminoglycoside antibiotic ( e . g . , In some aspects , identification and monitoring of bacterial tobramycin , gentamicin , kanamycin , amikacin , dibekacin , infection involves one or more of ( 1 ) nucleic acid detection sisomicin , netilmicin , neomycin B , neomycin C , neomycin methods , ( 2 ) serological detection methods, i . e ., conven E (paromomycin ), streptomycin ), a tetracycline antibiotic tional immunoassay, ( 3 ) culture methods, and ( 4 ) biochemi ( e . g . , tetracycline , chlortetracycline , oxytetracycline , deme- 55 cal methods . Such methods may be qualitative or quantita clocycline, lymecycline ,meclocycline , methacycline, mino tive. cycline , rolitetracycline , doxycyline ), or a ß - lactam antibi- Nucleic acid probes may be designed based on publicly otic ( e . g ., carbapenem , penicillin derivative (penam ) , available bacterial nucleic acid sequences , and used to detect cephalosporin (cephem ) , monobactam ) . target genes or metabolites (i . e. , toxins ) indicative of bac In particular embodiments , the antisense oligomer that 60 terial infection , which may be specific to a particular bac increases the sensitivity or reduces the MIC is targeted terial type , e .g . , a particular species or strain , or common to against infA , the bacterium is Escherichia coli or Acineto - more than one species or type of bacteria ( i. e. , Gram positive bacter baumannii that comprises or expresses InfA , and the or Gram negative bacteria ). Nucleic amplification tests ( e. g ., antimicrobial agent is an aminoglycoside antibiotic ( e . g . , PCR ) may also be used in such detection methods . tobramycin , gentamicin , kanamycin , amikacin , dibekacin , 65 Serological identification may be accomplished using a sisomicin , netilmicin , neomycin B , neomycin C , neomycin bacterial sample or culture isolated from a biological speci E ( paromomycin ), streptomycin ) , a tetracycline antibiotic men , e . g ., stool, urine , cerebrospinal fluid , blood , etc . Immu US 10 , 391 , 098 B2 73 74 noassay for the detection of bacteria is generally carried out mM NaCl and plated on tryptic - soy agar w / 5 % sheep blood by methods routinely employed by those of skill in the art, plate (Remel , Lenexa , Kans. ) to verify the starting concen e . g. , ELISA or Western blot. In addition , monoclonal anti tration . For minimal nutrient conditions , E . coli was incu bodies specific to particular bacterial strains or species are bated in MOPS minimal medium (Neidhart et al. , J Bacteriol often commercially available . 5 119 :736 -747 , 1974 ) , and Acinetobacter was performed in Culture methods may be used to isolate and identify AB minimal media ( Agrobacterium minimal media ) with 10 particular types of bacteria , by employing techniques includ - mM citrate ( Clark , J Mol Biol 23 : 99 - 112 , 1967 ) . ing, but not limited to , aerobic versus anaerobic culture , Minimal Inhibitory Concentration Assays . Minimal growth and morphology under various culture conditions. inhibitory concentration (MIC ) assays were performed in Exemplary biochemical tests include Gram stain (Gram , 10 Mueller Hinton II medium using the microdilution method 1884 ; Gram positive bacteria stain dark blue , and Gram as described by the Clinical and Laboratory Standards negative stain red ), enzymatic analyses , and phage typing Institute (CLSI ) . Optical density (OD ) of cultures was read It will be understood that the exact nature of such diag - in a microplate spectrophotometer at 595 -600 nm . After nostic , and quantitative tests as well as other physiological 18 - 20 hours of aerobic growth (200 - 250 rpm ) at 37° C . , 100 factors indicative of bacterial infection will vary dependent 15 ul cultures with an OD of < 0 . 06 were scored as no growth . upon the bacterial target, the condition being treated and Antibiotics were purchased from Sigma Chemical Co . ( St. whether the treatment is prophylactic or therapeutic . Louis , Mo. , USA ) . In cases where the subject has been diagnosed as having Minimal Bactericidal Concentration Assays . Minimum a particular type of bacterial infection , the status of the Bactericidal Concentration (MBC ) assays were performed bacterial infection is also monitored using diagnostic tech - 20 as MICs , with aliquots taken at noted time points , diluted in niques typically used by those of skill in the art to monitor 150 mM NaCl and plated . Plates were incubated for 18 the particular type of bacterial infection under treatment. hours and colonies were enumerated . The IC7 , of a PPMO The PMO or PPMO treatment regimen may be adjusted was defined as the MIC value in 75 % of the strains tested . ( dose , frequency, route , etc .) , as indicated , based on the Transmission election microscopy ( TEM ). At the speci results of immunoassays , other biochemical tests and physi - 25 fied time points the bacterial samples were centrifuged , ological examination of the subject under treatment . washed with Hank ' s Balanced Salt Solution (HBSS - ) (Life From the foregoing , it will be appreciated how various Technologies , Gibco , Grand Island , N . Y . , USA ) , resus objects and features of the present disclosure are met . The pended in 1 / 2 Karnovsky ' s fixative ( 4 % Paraformaldehyde , method provides an improvement in therapy against bacte - 2 . 5 % Glutaraldehyde and 0 . 1M Sodium Phosphate Buffer ) , rial infection , for example , multi - drug resistant (MDR ) 30 and stored at 4° C . until processing by the Electron Micros bacteria , using anti -acpP antisense oligomers to achieve copy Core Facility UT Southwestern Medical Center, Dal enhanced cell uptake and anti -bacterial action . As a result, las , Tex . The TEM grids were examined and images were drug therapy is more effective and less expensive , both in captured on a FEI TecnaiG2 Spirit Biotwin microsope (FEI terms of cost and amount of compound required . Company , Hillsboro , Oreg ., USA ). One exemplary aspect is that compounds effective against 35 Synergy Studies . MICswere performed with a PPMO and virtually any pathogenic bacterial can be readily designed a classic antibiotic ( as indicated ) in a 96 well plate . The and tested , e. g ., for rapid response against new drug - resis PPMO was diluted first horizontally down the plate and the tant strains . classic antibiotic was diluted across the rows of the plate . The following examples are intended to illustrate but not Inhibition was determined by visual observation and OD600 to limit the disclosure . Each of the patent and non -patent 40 ( see Berenbaum et al. , J Antimicrob Chemother 12 :555 - 563 , references referred to herein is incorporated by reference in 1983 ; and Berenbaum , J Infect Dis 137 : 122 - 130 , 1978 ) . For its entirety . determination of colony counts , aliquots of the target organ isms were incubated with the desired concentration of EXAMPLES PPMO , traditional antibiotic or combination of both and 45 grown at 37° C . , with centrifugation at 250 rpm . Cultures Materials and Methods were diluted in 150 mM NaCl at 0 and 24 hour time points , Peptide - Conjugated Phosphorodiamidate Morpholino plated and then colonies were enumerated . Synergy was Oligomers . PPMOs were synthesized and purified at Sarepta quantified using the method of isoboles as described (see Therapeutics Inc . (Cambridge , Mass . , USA ) as previously Tallarida , Genes Cancer. 2 : 1003 - 8 , 2011 ) : (Effective con described ( Tilley et al. , Antimicrob Agents Chemother 50 centration of antibiotic /MIC of antibiotic ) + ( Effective con 50 : 2789 - 2796 , 2006 ) . Lyophilized PPMOs were dissolved centration of PPMO /MIC of PPMO ). Calculated values less in ultrapure water and sterile - filtered . PPMO peptides were than 1 indicated synergy . attached to either the 5 ' or 3 ' end of the oligomer sequence Graphical Software . Standard deviation and graphical as indicated . analysis was performed on GraphPad Prism®6 software Bacteria . Bacterial strains were obtained through the 55 (GraphPad Software, Inc. , San Diego , Calif ., USA ) . clinicalmicrobiology lab at UT Southwestern unless other wise noted . Strains also included those purchased through Example 1 ATCC , the E . coli Genetic Stock Center ( Yale University ), or received from collaborators . They included both genome Activity of PPMO Targeted Against acpP of E . coli sequenced isolates as well as clinical isolates with varying 60 levels of antibiotic resistance . Single colonies were grown A cell -penetrating peptide -conjugated phosphorodiami aerobically to stationary phase ( 1 - 2 mLs at 37° C ., 250 rpm ) date morpholino oligomer ( PPMO ) targeted against the E . in Mueller Hinton cation -adjusted (MH II ) broth (Becton - coli acpP gene was prepared and its efficacy was evaluated Dickinson Difco BBL , Franklin Lakes , N . J ., USA ) . The in vitro against a multi - drug - resistant strain of E . coli OD600 was taken and a working stock of 5x10° was 65 (A15070834 ) . generated based on the individual strain ' s colony forming The acpP - targeted PPMO # 2 has the following sequence : units ( CFU )/ mL /OD . Working stocks were diluted in 150 5 '- CTTCGATAGTG - 3 ' (SEQ ID NO : 1 ) . The PPMO was US 10 , 391 ,098 B2 75 76 conjugated at its 3 '- end to the C - terminal beta - alanine of the multi - drug - resistant strain of Acinetobacter baumannii (RXR ) XB (SEQ ID NO :59 ) . ( AYE ) by about 6 - logs relative to scramble PPMO control, The acpP - targeted PPMO ( 1 uM ) was added to bacterial but in combination with colistin also synergistically reduced cultures either alone or in combination with tobramycin (2 bacterial growth to undetectable levels (an additional ~ 3 ug /ml ) . PPMO scramble control ( 1 uM ), peptide control ( 1 5 logs relative to the acpP - targeted PPMO alone ). Colistin and uM ) , and tobramycin ( 2 ug /ml ) were added to bacterial control PPMOs ( either alone or in combination ) did not have cultures separately or in various combinations . Colony - this significant of an effect on bacterial growth . forming units (CFU ) were counted at 24 hours . FIG . 4 shows that the acpP - targeted PPMO # 7 not only As shown in FIG . 2A , the acpP - targeted PPMO # 2 not reduced bacterial growth ( colony - forming units; CFU ) of only reduced bacterial growth ( colony - forming units ; CFUS ) 10 the multi - drug -resistant strain of Acinetobacter baumannii of the multi - drug -resistant strain of E . coli by about 1 - log ( AYE ) by about 5 - logs relative to scramble PPMO control, relative to scramble PPMO and peptide controls , but in but in combination with meropenem also synergistically combination with tobramycin also synergistically reduced reduced bacterial growth by an additional ~ 2 - logs relative to bacterial growth by over 4 -logs relative to controls . the acpP - targeted PPMO alone. Meropenem and control Tobramycin and control PPMOs ( either alone or in combi- 15 PPMOs (either alone or in combination ) did not have this nation ) had no significant effect on bacterial growth . significant of an effect on bacterial growth . Example 2 FIG . 5 shows that the acpP - targeted PPMO # 7 not only reduced bacterial growth ( colony - forming units ; CFUS) of Effect of acpP - Targeted PPMO on MIC of 20 the multi - drug - resistant strain of Acinetobacter baumannii Tobramycin ( AYE ) by about 6 - logs relative to scramble PPMO control, but in combination with tobramycin also synergistically A PPMO was tested for its effects on the minimum reduced bacterial by an additional ~ 1 - log relative to the inhibitory concentration (MIC ) of tobramycin against a acpP -targeted PPMO alone. Tobramycin and control PPMOs multi - drug - resistant strain of E . coli ( A15070834 ) . 25 (either alone or in combination ) did not have this significant The acpP - targeted PPMO # 1 has the following sequence : of an effect on bacterial growth . 5 ' -CTTCGATAGTG - 3 ' (SEQ ID NO : 1 ) . The PPMO was conjugated at its 5 ' - end to the C - terminal beta - alanine of Example 4 (RXR ) XB (SEQ ID NO :59 ). The MIC of the tobramycin was measured using the 30 Effect of acpP - Targeted PPMO on MIC of microdilution method of the Clinical Laboratory Standards Antibiotics Institute in a 96 -well microtiter plate format . Multiple , identical dilution series of tobramycin were included on The PPMO # 7 from Example 3 was tested for its effects on each microtiter plate . In each dilution series of tobramycin , the minimum inhibitory concentration (MIC ) of colistin , a fixed amount of PPMO was added . Each dilution series of 35 meropenem , and tobramycin against multi - drug -resistant antibiotic included a different concentration of PPMO . The strains of Acinetobacter baumannii ( AYE ) and E . coli results are shown in FIGS. 2B - 2C . AIS070834 . FIG . 2B ( linear scale ) and FIG . 2C ( log scale ) show that The MIC of the antibiotics colistin , meropenem , and increasing amounts of acpP -targeted PPMO # 1 significantly tobramycin was measured using the microdilution method of decreased the minimum inhibitory concentration (MIC ) of 40 the Clinical Laboratory Standards Institute in a 96 -well tobramycin against a multi - drug -resistant strain of E . coli in microtiter plate format . Multiple, identical dilution series of a concentration - dependent manner. each antibiotic were included on each microtiter plate . In each dilution series of antibiotic , a fixed amount of PPMO Example 3 was added . Each dilution series of antibiotic included a 45 different concentration of PPMO . The results are shown in Activity of PPMO Targeted Against acpP of A . FIGS . 3 - 8 . baumannii FIG . 6 shows that increasing amounts of the acpP - targeted PPMO # 7 significantly decreased theMIC of colistin against A cell -penetrating peptide - conjugated phosphorodiami- the multi - drug -resistant strain of Acinetobacter baumannii date morpholino oligomer (PPMO ) targeted against the A . 50 (AYE ) in a concentration -dependent manner . In the presence baumannii acpP gene was prepared and its efficacy was of 2 uM of PPMO , the MIC of colistin decreased from 1 evaluated in vitro against a multi -drug - resistant strain of A . ug/ mL to 0 . 25 ug/ mL (see also FIG . 3) , and synergy was baumannii (AYE ). 0 .75 . The acpP - targeted PPMO # 7 has the following sequence : FIG . 7 shows that increasing amounts of the acpP -targeted 5 ' - ATATCGCTCAC - 3 ' ( SEO ID NO : 2 ). The PPMO was 55 PPMO # 7 also significantly decreased the MIC of mero conjugated at its 3 '- end to the C -terminal beta -alanine of penem against the multi - drug -resistant strain of Acineto (RXR ) XB (SEQ ID NO :59 ) . bacter baumannii (AYE ) in a concentration - dependentman The acpP -targeted PPMO # 7 ( 2 uM ) was added to bacte ner. In the absence of PPMO theMIC of the meropenem was rial cultures either alone or in combination with colistin 2 ug/ mL , and this was reduced to 0 .25 ug /mL when 4 uM ( 0 . 25 ug /ml ) , meropenem ( 0 . 5 ug /ml ) , or tobramycin ( 2 60 PPMO was present (see also FIG . 4 ) . Synergy between ug /ml ) . PPMO scramble control ( 2 UM ) , colistin ( 0 . 25 meropenem and the acpP - targeted PPMO was 0 .75 . ug/ ml ) , meropenem (0 . 5 ug/ ml ) , and tobramycin ( 2 ug /ml ) This effect was not limited to antibiotics that affected were added to bacterial cultures separately or in various bacterial membrane structure as synergy was also seen with combinations . Colony - forming units (CFU ) were counted the aminoglycoside tobramycin . FIG . 8 shows that increas at 24 hours . The results are shown in FIGS. 3 - 5 . 65 ing amounts of the acpP - targeted PPMO # 7 significantly FIG . 3 shows that the acpP - targeted PPMO # 7 not only decreased the MIC of tobramycin against the multi -drug reduced bacterial growth ( colony - forming units ; CFUs) of resistant strain of Acinetobacter baumannii (AYE ) in acon US 10 , 391 , 098 B2 77 78 centration -dependent manner . In the absence of PPMO , the MICs were performed in both rich and minimal media for MIC of tobramycin was 64 ug /mL . The MUC was reduced both genera of bacteria . Panels of E . coli ( ~ 22 strains ) and to 2 ug/ mL in the presence of 4 uM PPMO ( see also FIG . 5 ) . Acinetobacter spp . (~ 34 strains ) comprised both drug -sen The synergy between tobramycin and the acpP -targeted sitive and multidrug -resistant strains, as shown in Table El PPMO was 0 .625 . below . TABLE E1 Pathogen Strain Characteristics Source E . coli W3110 non - pathogenic , K - 12 E . coli Genetics Stock Center, New Haven , CT E . coli SMS- 3 - 5 MDR ATCC E . coli CVB - 1 NDM1 producer G . Rossolini, University of Siena , Italy E . coli BCT- B -036NDM - 1 NDM1 producer P . Nordmann , Hopital de Bicetre , Paris E . coli NDM1- E NDM1 producer S . Poutanen , Mt. Sinai Hospital , Toronto E . coli BAA - 196 ESBL , TEM - 10 ATCC E . coli BAA - 200 MDR , SVH - 4 ATCC E . coli BAA - 202 ESBL , ceftazidine resist . ATCC E . coli ATCC700928 UTI , genome sequenced ATCC E . coli ATCC 25922 Control strain for MIC ATCC suscept . E . coli E2348 /69 EPEC J . Kaper, University of Maryland , Baltimore, MD E . coli 1001728 NDM1 producer J. K . Rasheed , CDC , Atlanta , GA E . coli 1101851 NDM1 producer J . K . Rasheed , CDC , Atlanta , GA E . coli A15070834 NDM1 producer J . K . Rasheed , CDC , Atlanta , GA E . coli A1071077 NDM1 producer J . K . Rasheed , CDC , Atlanta , GA A . baumannii BAA - 1710 MDR , genome sequenced ATCC A . baumannii BAA - 1709 Genome sequenced ATCC A . baumannii AB0057 MDR Todd Hoopman , UTSW A . baumannii ATCC 17978 Genome sequenced ATCC A . baumannii ATCC 17961 ATCC A . baumannii ATCC 17906 ATCC A . baumannii AYE (PNDM - 1 ) AYE with NDM - 1 Bruce Geller, OSU A . baumannii BCT- 13 -026NDM - 1 NDM1 producer P . Nordmann , Hopital de Bicetre , Paris A . baumannii ATCC 19606 Genome sequenced ATCC A . iwoffii ATCC 17976 Genome sequenced ATCC A . baumannii HUMC - 1 Brad Spellberg , USC

For all antibiotics tested , there was a corresponding list of all PPMOs used and their sequences are provided reduction in CFU /ml of at least 1 - log when PPMO was ( Tables 3A - B ) . PPMOs were designed against known or combined with the antibiotic compared to either the PPMO putative essential bacterial genes in a variety of pathways or antibiotic alone . The scrambled PPMO at 8 uM or less , 45 including fatty acid , lipopolysaccharide or peptidoglycan showed no activity alone or synergy with the antibiotics biosynthesis . Regardless of themedia used , PPMOs targeted tested . to the acyl carrier protein (AcpP ) showed the best in vitro Similar synergy between the acpP (PPMO # 1) and the inhibition in both E . coli and Acinetobacter. In MHII , 6 PPMOs were the most active in E . coli with an IC 5 of 4 uM same three antibiotics was seen in the E . coli strain 50 or less ( FIG . 13A ) . Five of the 6 were all targeted to acpP A15070834 (see FIGS. 12A - 12F ) . FIGS. 12A - 12B show the with the differences related to various peptide attachments or results for colistin , FIGS. 12C - 12D show the results for position on the target. The acpP PPMOs had IC75 MICs that meropenem , and FIGS . 12E - 12F show the results for ranged from 1 to 4 uM . The other active PPMO targeted tobramycin . In all instances , the acpP -targeted PPMO sig murA and had an IC , MICs of 4 uM . nificantly reduced the MIC of the tested antibiotic . The freeset 55 The determination of whether a bacterial gene is essential peptide ( RXR )4XB was also tested and by itself had an can vary based on the media in which the bacteria are grown . unmeasurable MIC (data not shown ) . Fifteen E . coli strains were screened in MOPS minimal media ( FIG . 13B ) . This screen increased the number of Example 5 PPMOs with IC75 MICs of 4 uM or less to 10 . Six of the ten 60 represented the PPMOs that were found to be effective in PPMOs Targeting Essential Genes Inhibit Growth MHII media although the MIC values improved with the of Acinetobacter Spp . and E . coli In Vitro most potent acpP PPMO having IC75 MICs of 0 . 5 UM . Additional potent PPMOs were identified and included the PPMOs targeted against acpP were tested for the ability to gene targets : rpmB (a recombinant ribosomal protein gene; inhibit the growth of Acinetobacter spp . and E . coli in vitro . 65 IC75 MICs of 0 . 5 and 1 uM ) , adk (an adenylate kinase gene ; Bacterial strains and culture and assay conditions are IC75 MIC of 2 uM ) and infA ( a transcription antiterminator described in the Materials and Methods section above . gene ; IC75 MIC of 4 uM ) . US 10 , 391 , 098 B2 79 80 In Acinetobacter , acpP PPMOs # 6 and # 7 had IC75 MICS E . coli, including MDR strains, are bactericidal at clinically of 2 to 4 uM , respectively, in nutrient rich MHII media . relevant concentrations (e . g ., IC75 of 4 uM or less ) . These These PPMOs differed only in the positioning of the same data also show PPMOs targeted against the acpP and other peptide on the 5 ' (PPMO # 6 ) or 3 ' (PPMO # 7 ) end of the genes showed synergy with classic antibiotics tobramycin , oligomer ( FIG . 13C ) . As was seen in E . coli , Acinetobacter 5 meropenem and colistin , and restored the efficacy of those tested in minimal media increased the number of PPMOs antibiotics in MDR strains of Acinetobacter and Escheri that showed in vitro activity . Four additional acpP PPMOs chia . PPMOs could therefore be used either alone or syn were found to have activity along with the two that were ergistically with traditional antibiotics . When sub - inhibitory active in MHII with IC , MICs ranginganging fromfrom 2 toto 44 uM . concentrations of the antibiotic were used in combination Further gene targets were identified in AB minimal media 10 with a PPMO viability or growth of A . baumannii and E . coli including ftsZ (Cell division Z ring ) with an IC75 of 4 uM was significantly reduced . For some antibiotics, such as and accA ( carboxyltransferase alpha subunit of acetyl Coen - colistin , the combination with PPMOs led to reduced viabil zyme A carboxylase ) with an IC75 of 4 UM ( FIG . 13D ) . ity by > 3 logs. The enhanced activity of PPMOs in minimal media was maintained in MDR strains as well , with E . coli W3110 and 15 Example 6 A . baumannii AYE and 0057 having MIC values that were 1 - 2 fold lower when compared to rich media ( see FIGS. Effect of acpP - Targeted PPMO on Acinetobacter 13A -D ; FIGS . 9A -9C ; FIGS. 10A - 10C ; and FIGS . 12A Cell Wall 12C ) . FIG . 9A shows the results for E . coli W3110 challenged 20 Transmission electron microscopy ( TEM ) was used to with acpP ( PPMO # 1 ) , acpP ( PPMO # 2 ) , acpP (PPMO # 3 ) , study the morphological alterations to Acinetobacter follow acpP (PPMO # 4 ), acpP (PPMO # 5 ) , murA (PPMO # 24 ) , and ing challenge with acpP - targeted PPMOS. A . baumannii Scramble ( Scr ) controls . FIGS . 9B - 9C respectively shows AYE was grown in nutrient rich MHII media (Mueller the results for A . baumannii AYE ( 9B ) and A . baumannii Hinton cation -adjusted broth ; Becton -Dickinson Difco 0057 (9C ) challenged with acpP (PPMO # 7) , acpP 25 BBL , Franklin Lakes, N .J ., USA ) in the presence or absence (PPMO # 8 ) , acpP (PPMO # 13 ) , acpP (PPMO # 14 ), ftsZ of acpP PPMO (PPMO # 7 ) or Scramble (Scr ) PPMO control ( PPMO # 33 ) , ftsz ( PPMO # 34 ), rpmB (PPMO # 29 ) , and at a concentration of 40 uM . FIGS . 11A and 11B show A . Scramble (Scr ) controls . baumannii AYE at 0 hour with an intact cell wall and PPMOs were also bactericidal in multidrug - resistant cytoplasmic space in the absence of any PPMO . This was (MDR ) strains, as measured by kinetic MBC assays . The 30 also true of AYE in the presence of Scr PPMO (FIGS . MDR strains E . coli 1101851, A . baumannii AYE , and A . 11C - 11D ) and acpP PPMO (FIGS . 11E - 11F ) at 0 hour. After baumannii 0057 were grown in the presence or absence of 6 hours of incubation , cells incubated with Scr PPMO different concentrations of PPMOs and samples were plated (FIGS . 111- 11J) were indistinguishable from those of the at various time points to determine the amount of viable untreated samples (FIGS . 114 - 11H ). In contrast, after 6 bacteria present (MBC assays , as described above ). The 35 hours of incubation with the acpP -targeted PPMO , cell wall results are shown in FIGS . 10A - 10C . Despite being multi - disruption was observed ( FIGS. 11K - 11L ). This disruption drug -resistant , PPMOs demonstrated both time and concen was seen as early as 3 hours (data not shown ). These results tration dependent killing . By two hours , PPMOs at a con - suggest that the reduction in Acinetobacter cell viability in centration of 1 to 2 um decreased viability by greater than the presence of Acp PPMO is due at least in part to the 4 logs in E . coli 1101851 (FIG . 10A ) . By eight hours , all 40 ability of the PPMO to cause cell wall damage . concentrations tested of 0 .0625 uM or greater were bacte ricidal and below the limit of detection . Example 7 PPMOs also demonstrated both time and concentration dependent killing in A . baumannii strains AYE and AB0057 , PPMOs are Synergistic in Combination with although with higher concentrations of PPMO and at a 45 Traditional Antibiotics slower rate of kill than in E . coli . At 24 hours, the acpP targeted PPMO ( PPMO # 7 ) was bactericidal and below the To determine whether the combination of active PPMOS limit of detection at PPMO concentrations of > 4 uM in both and traditional antibiotics were additive or synergistic in strains tested (FIGS . 10B - 10C ) . A PPMO with a scrambled their effects , checkerboard MIC assays were performed . The oligo sequence linked to the same peptide (Scr - (RXR ) 4 ) had 50 multidrug resistant E . coli AIS070834 was incubated with no effect on any strain ( FIG . 10A - 10C ) . increasing concentrations of ftsZ PPMO (PPMO # 46 ) or There was also synergy between acpP - targeted PPMOs scrambled (Scr ) PPMO ( Scr- 1 ) and increasing concentra and three different antibiotics against the multidrug - resistant tions of either colistin , meropenem or tobramycin . E . coli strain AIS070834 (see FIGS. 12A - 12F ) . The MIC of In the presence of 2 uM of ftsZ PPMO , the MIC of colistin colistin , meropenem , and tobramycin was measured with 55 decreased from 0 .5 ug /mL to 0 .05 ug/ mL (FIG . 14A ) . In various concentrations of acpP - targeted PPMO ( PPMO # 1 ) addition , the MIC of meropenem decreased with increasing or scrambled ( Ser ) control PPMO . Viable cells were counted concentrations of ftsZ PPMO . In the absence of ftsZ PPMO in 24 -hour cultures with antibiotic alone , PPMO alone, or in the MIC of meropenem was 12 ug /mL , and this was reduced combination thereof. FIGS . 12A - 12B show the results for to 2 ug /mL when 4 um ftsZ PPMO was present ( FIG . 14C ) . colistin , FIGS . 12C - 12D show the results for meropenem , 60 This effect was not limited to antibiotics that affected and FIGS . 12E - 12F show the results for tobramycin . In all bacterial membrane structure as synergy was also seen with instances , the acpP - targeted PPMO significantly reduced the the aminoglycoside tobramycin . In the absence of ftsz MIC of the tested antibiotics . The free peptide (RXR )4XB PPMO , the MIC of tobramycin was 270 ug /mL . This was was also tested and by itself had an unmeasurable MIC ( data reduced to almost an undetectable level in the presence of 2 not shown ) . 65 UM PPMO (FIG . 14E ) . For all antibiotics tested , there was Overall , these data show , inter alia , that PPMOs targeted a corresponding reduction in CFU /ml of at least 4 logs when against the acpP and other genes of Acinetobacter spp . and ftsZ PPMO was combined with the antibiotic compared to US 10 , 391 , 098 B2 81 82 either the PPMO or antibiotic alone ( FIG . 14B , FIG . 14D , strains. While SMS- 3 - 5 is resistant to chloramphenicol FIG . 14F ) . The scrambled PPMO at 32 uM or less, or free (MIC > 512 ug /mL ), when incubated with increasing con peptide (RXR ) XB at 32 uM or less, showed no activity centrations of a cmlA PPMO (PPMO # 67 ) , the MIC also alone or synergy with the antibiotics tested . progressively decreases in a dose -dependent fashion (FIG . 5 15B ) . Example 8 To see whether blocking antibiotic resistance genes with PPMOs could be effective in other genera , A . baumannii PPMOs Targeted to Nonessential AYE was incubated with a PPMO against adeA (PPMO # 65) , Antibiotic -resistance Genes Restore Susceptibility which encodes a component of the AdeABC RND - type of MDR Strains to Traditional Antibiotics 10 multidrug efflux pump . AdeABC confers resistance to a Modulating antibiotic resistance with PPMOs could be an variety of antibiotics including aminoglycosides . AYE was alternative therapeutic strategy. As proof of concept, PPMOs treated with varying concentrations of tobramycin and adeA were designed to target specific , non -essential antibiotic PPMO in both MHII and AB Minimal Media . While resistance genes. blaT is part of the TEM beta - lactamase 15 tobramycin alone had an MIC of 64 ug /mL in both media , family and is found in environmental strains of E . coli , like increasing concentrations of the adeA PPMO reduce the SMS - 3 - 5 . While SMS- 3 -5 is resistant to the beta - lactam MIC of tobramycin significantly (FIG . 16A , FIG . 16B ). ampicillin (MIC > 1024 ug /mL ) , when incubated with With 8 uM adeA PPMO , the MIC of tobramycin was increasing concentrations of a blaT PPMO ( PPMO # 66 ) , the reduced to 4 ug/ mL and 1 ug /mL in MHII and minimal MIC progressively decreased ( FIG . 15A ) . cmlA is an amino media , respectively . A scrambled PPMO at 8 uM had no glycoside resistance gene found in environmental E . coli effect on the MIC of tobramycin .

SEQUENCE LISTING

< 160 > NUMBER OF SEQ ID NOS : 71 < 210 > SEQ ID NO 1 < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : acpp targeting sequence < 400 > SEQUENCE : 1 cttcgatagt g 11

< 210 > SEQ ID NO 2 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : acpP targeting sequence < 400 > SEQUENCE : 2 atatcgctca C 11

< 210 > SEQ ID NO 3 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 220 > FEATURE : < 223 > OTHER INFORMATION : acpp targeting sequence < 400 > SEQUENCE : 3 attctcctcat 11

< 210 > SEQ ID NO 4 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial ? 20 > FEATURE : < 223 > OTHER INFORMATION : acpp targeting sequence < 400 > SEQUENCE : 4

cacagga att C 11

< 210 > SEQ ID NO 5 US 10 ,391 , 098 B2 83 84 - continued < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : acps targeting sequence < 400 > SEQUENCE : 5 ttgccattag c 11

< 210 > SEQ ID NO 6 < 211 > LENGTH : 17 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : acp - E targeting sequence < 400 > SEQUENCE : 6 ctgtagtgat ttcacca 17

< 210 > SEQ ID NO 7 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : fabA targeting sequence < 400 > SEQUENCE : 7 ttatctacca t 11

< 210 > SEQ ID NO 8 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : fabB targeting sequence < 400 > SEQUENCE : 8 gcacgtttca t 11

< 210 > SEQ ID NO 9 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : fabi targeting sequence < 400 > SEQUENCE : 9 agaaaacccat 11

< 210 > SEQ ID NO 10 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223NNHAHOWN > OTHER INFORMATION : gapA targeting sequence < 400 > SEQUENCE : 10 ttgatagtcat 11

< 210 > SEQ ID NO 11 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213??? > ORGANISM : artificial < 220 > FEATURE : < 223??? > OTHER INFORMATION : accA targeting sequence < 400 > SEQUENCE : 11 US 10 ,391 ,098 B2 85 86 - continued gcttttttca t 11

< 210 > SEQ ID NO 12 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : mura targeting sequence < 400 > SEQUENCE : 12 atccatttag t 11

< 210 > SEQ ID NO 13 < 211 > LENGTH : 11 TYPE : DNA ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : murA targeting sequence < 400 > SEQUENCE : 13 catttagttt g 11

< 210 > SEQ ID NO 14 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : mura targeting sequence < 400 > SEQUENCE : 14 aatttatcca t 11

< 210 > SEQ ID NO 15 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223??????WOWNP > OTHER INFORMATION : mura targeting sequence < 400 > SEQUENCE : 15 aaatttatcc a 11

< 210 > SEQ ID NO 16 < 211 > LENGTH : 11 < 212 > TYPE : DNA ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : rpmB targeting sequence < 400 > SEQUENCE : 16 actcgggaca t 11

< 210 > SEQ ID NO 17 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : rpmB targeting sequence < 400 > SEQUENCE : 17 ctattctcca a 11

< 210 > SEQ ID NO 18 < 211 > LENGTH : 11 < 212 > TYPE : DNA US 10 ,391 , 098 B2 87 88 - continued < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : rpmB targeting sequence < 400 > SEQUENCE : 18 ggcagactcg g 11

< 210 > SEQ ID NO 19 V< 211 > LENGTH : 11 V< 212 > TYPE : DNA V< 213 > ORGANISM : artificial V< 220 > FEATURE : < 223 > OTHER INFORMATION : rpmB targeting sequence < 400 > SEQUENCE : 19 cttagacatg g 11

< 210 > SEQ ID NO 20 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 A> ORGANISM : artificial < 220 A> FEATURE : < 223 > OTHER INFORMATION : adk targeting sequence < 400 > SEQUENCE : 20 atgatacgca t 11

< 210 > SEQ ID NO 21 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : infA targeting sequence < 400 > SEQUENCE : 21 tctttggcca t 11

< 210 > SEQ ID NO 22 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : ftsz targeting sequence < 400 > SEQUENCE : 22 tcaaatgagg c 11

< 210 > SEQ ID NO 23 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : ftsz targeting sequence < 400 > SEQUENCE : 23 aatgaggccat 11

< 210 > SEQ ID NO 24 < 211 > LENGTH : 13 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : ftsz targeting sequence < 400 > SEQUENCE : 24 atagtttctc tcc 13 US 10 ,391 , 098 B2 89 - continued

< 210 > SEQ ID NO 25 < 211 > LENGTH : 11 < 212 > TYPE : DNA ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : rpoD targeting sequence < 400 > SEQUENCE : 25 tcatctttgc t 11

< 210 > SEQ ID NO 26 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : rpoD targeting sequence < 400 > SEQUENCE : 26 ttttgctcca t 11

< 210 > SEQ ID NO 27 < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial 20 > FEATURE : < NNN223 > OTHER INFORMATION : aroc targeting sequence < 400 > SEQUENCE : 27 ttccctgccat 11

< 210 > SEQ ID NO 28 < 211 > LENGTH : 11 < 212 > TYPE : DNA V 13 > ORGANISM : artificial

V 20 > FEATURE : < 223 > OTHER INFORMATION : aroc targeting sequence < 400 > SEQUENCE : 28 tttccagecat 11

< 210 > SEO ID NO 29 < 211 > LENGTH : 11 < 212 > TYPE : DNA 3 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : murf targeting sequence < 400 > SEQUENCE : 29 acgctaatca t 11

< 210 > SEO ID NO 30 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : 1pxC targeting sequence < 400 > SEQUENCE : 30 tgtttgatca t 11

< 210 > SEQ ID NO 31 < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial < 220 > FEATURE : US 10 ,391 , 098 B2 91 - continued < 223 > OTHER INFORMATION : kdtA targeting sequence < 400 > SEQUENCE : 31 aattcgagcat 11

< 210 > SEQ ID NO 32 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : boxa targeting sequence < 400 > SEQUENCE : 32 tgttaaagag C 11

< 210 > SEQ ID NO 33 < 211 > LENGTH : 15 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : rpoD - E targeting sequence < 400 > SEQUENCE : 33 cttgtaacca cacca 15

< 210 > SEQ ID NO 34 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : pryc targeting sequence < 400 > SEQUENCE : 34 ggtgcagtcat 11

< 210 > SEQ ID NO 35 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : pryA targeting sequence < 400 > SEQUENCE : 35 gacttaatca a 11

< 210 > SEQ ID NO 36 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : lgt targeting sequence < 400 > SEQUENCE : 36 ctactggtcat 11

< 210 > SEQ ID NO 37 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : fola targeting sequence < 400 > SEQUENCE : 37 cattgagatt t 11 US 10 ,391 , 098 B2 93 94 - continued < 210 > SEO ID NO 38 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 2 3 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : infB targeting sequence < 400 > SEQUENCE : 38 acatctgtcat 11

< 210 > SEO ID NO 39 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : nrda targeting sequence < 400 > SEQUENCE : 39 ttctgattca t 11

< 210 > SEQ ID NO 40 < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : nrdB targeting sequence < 400 > SEQUENCE : 40 gtatatgcca t 11

< 210 > SEQ ID NO 41 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : zipA targeting sequence < 400 > SEQUENCE : 41 tcctgcatcat 11

< 210 > SEQ ID NO 42 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : coaA targeting sequence < 400 > SEQUENCE : 42 atatacctca t 11

< 210 > SEQ ID NO 43 < 211 > LENGTH : 17 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : gyrA - E targeting sequence < 400 > SEQUENCE : 43 gttaccctga ccgacca 17

< 210 > SEQ ID NO 44 < 211 > LENGTH : 16 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : V 223 > OTHER INFORMATION : qyrA - E targeting sequence US 10 ,391 ,098 B2 95 96 - continued < 400 > SEQUENCE : 44 gttaccctga ccacca 16

< 210 > SEQ ID NO 45 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial V 20 > FEATURE : < 223 > OTHER INFORMATION : mrdA targeting sequence < 400 > SEQUENCE : 45 tgtttcatac g 11

V< 210 > SEQ ID NO 46 V< 211 > LENGTH : 11 V< 212 > TYPE : DNA V< 213 > ORGANISM : artificial < 220 > FEATURE : < 223> OTHER INFORMATION : 1pxB targeting sequence < 400 > SEQUENCE : 46 ggtttgccaa g 11

< 210 > SEQ ID NO 47 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : 1pxC targeting sequence < 400 > SEQUENCE : 47 tgtttcacca t 11

< 210 > SEQ ID NO 48 < 211 > LENGTH : 11 A TYPE : DNA A ORGANISM : artificial NNNNN A FEATURE : < 223 > OTHER INFORMATION : kdta targeting sequence 20 > FEATURE : < 221 > NAME /KEY : misc _ feature < 222 > LOCATION : ( 1 ) . . ( 1 ) < 223 > OTHER INFORMATION : I < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 222 > LOCATION : ( 2 ) . . ( 2 ) < 223 > OTHER INFORMATION : I < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 222 > LOCATION : ( 3 ) . . ( 3 ) < 223 > OTHER INFORMATION : I 20 > FEATURE : < 221 > NAME / KEY : misc _ feature ? 2 > LOCATION : ( 4 ) . . ( 4 ) < 223 > OTHER INFORMATION : I < 400 > SEQUENCE : 48 nnnntcgcca a 11

< 210 > SEQ ID NO 49 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : boxA targeting sequence < 400 > SEQUENCE : 49 ctcttaatga t 11 US 10 ,391 , 098 B2 97 98 - continued

< 210 > SEQ ID NO 50 < 211 > LENGTH : 11 < 212 > TYPE : DNA ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : boxC targeting sequence < 400 > SEQUENCE : 50 atocacacaa g 11

< 210 > SEQ ID NO 51 < 211 > LENGTH : 15 < 212 > TYPE : DNA < 213 > ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : rpoD - E targeting sequence < 400 > SEQUENCE : 51 tccaccaaqt cacca 15

< 210 > SEQ ID NO 52 < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial 20 > FEATURE : < 223MOM > OTHER INFORMATION : pryc targeting sequence < 400 > SEQUENCE : 52 agagttcaag g 11

< 210 > SEQ ID NO 53 < 211 > LENGTH : 11 < 212 > TYPE : DNA ORGANISM : artificial 20 > FEATURE : < 223 > OTHER INFORMATION : cara targeting sequence < 400 > SEQUENCE : 53 ggtgctcaaa c 11

< 210 > SEO ID NO 54 < 211 > LENGTH : 11 < 212 > TYPE : DNA 3 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : adeA targeting sequence < 400 > SEQUENCE : 54 atactgtcca a 11

< 210 > SEO ID NO 55 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : blat targeting sequence < 400 > SEQUENCE : 55 ctcttccttt t 11

< 210 > SEQ ID NO 56 < 211 > LENGTH : 11 < 212 > TYPE : DNA 13 > ORGANISM : artificial < 220 > FEATURE : US 10 ,391 , 098 B2 99 100 - continued < 223 > OTHER INFORMATION : ?ml targeting sequence < 400 > SEQUENCE : 56 tccttctgat t 11

< 210 > SEQ ID NO 57 < 211 > LENGTH : 12 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : < 221 > NAME /KEY : MOD _ RES < 222 > LOCATION : ( 2 ) . . ( 2 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME /KEY : MOD _ RES < 222 > LOCATION : ( 5 ) . . ( 5 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 8 ) . . ( 8 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 11 ) . . ( 11 ) < 223 > OTHER INFORMATION : Acp < 400 > SEQUENCE : 57 Arg Xaa Arg Arg Xaa Arg Arg Xaa Arg Arg Xaa Arg 10

< 210 > SEQ ID NO 58 < 211 > LENGTH : 10 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 400 > SEQUENCE : 58 Arg Phe Phe Arg Phe Phe Arg Phe Phe Arg 10

< 210 > SEQ ID NO 59 < 211 > LENGTH : 14 < 212 > TYPE : PRT A 13 > ORGANISM : artificial < 2 20 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 2 ) . . ( 2 ) < 223 > OTHER INFORMATION : Acp A? 20 > FEATURE : < 221 > NAME / KEY : MOD _RES < 222 > LOCATION : ( 5 ) . . ( 5 ) < 223 > OTHER INFORMATION : Acp A? 20 > FEATURE : < 221 > NAME / KEY : MOD _RES < 222 > LOCATION : ( 8 ) . . ( 8 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME /KEY : MOD _ RES < 222 > LOCATION : ( 11 ) . . ( 11 ) < 223 > OTHER INFORMATION : Acp 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 13 ) . . ( 13 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME /KEY : MOD _ RES < 222 > LOCATION : ( 14 ) . . ( 14 ) < 223 > OTHER INFORMATION : bAla US 10 ,391 , 098 B2 101 102 - continued < 400 > SEQUENCE : 59 Arg Xaa Arg Arg Xaa Arg Arg Xaa Arg Arg Xaa Arg Xaa Xaa 10

< 210 > SEQ ID NO 60 < 211 > LENGTH : 12 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : < 221 > NAME /KEY : MOD _ RES < 222 > LOCATION : ( 11 ) . . ( 11 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 12 ) . . ( 12 ) < 223 > OTHER INFORMATION : bAla < 400 > SEQUENCE : 60 Arg Phe Phe Arg Phe Phe Arg Phe Phe Arg Xaa Xaa 10

< 210 > SEQ ID NO 61 < 211 > LENGTH : 12 < 212 > TYPE : PRT < 213????? > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 400 > SEQUENCE : 61 Arg Phe Arg Arg Phe Arg Arg Phe Arg Arg Phe Arg 10

< 210 > SEQ ID NO 62 < 211 > LENGTH : 12 < 212 > TYPE : PRT ORGANISM : artificial FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 400 > SEQUENCE : 62 Arg Tyr Arg Arg Tyr Arg Arg Tyr Arg Arg Tyr Arg 10

< 210 > SEQ ID NO 63 V 11 > LENGTH : 12 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 400 > SEQUENCE : 63 Arg Gly Arg Arg Gly Arg Arg Gly Arg Arg Gly Arg 10

< 210 > SEQ ID NO 64 < 211 > LENGTH : 14 < 212 > TYPE : PRT < 213 > ORGANISM : artificial ?N FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 13 ) . . ( 13 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES US 10 ,391 , 098 B2 103 104 - continued < 222 > LOCATION : ( 14 ) . . ( 14 ) < 223 > OTHER INFORMATION : bAla < 400 > SEQUENCE : 64 Arg Phe Arg Arg Phe Arg Arg Phe Arg Arg Phe Arg Xaa Xaa 10

< 210 > SEQ ID NO 65 < 211 > LENGTH : 14 < 212 > TYPE : PRT < 213 > ORGANISM : artificial FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : < 221 > NAME / KEY : MOD _RES < 222 > LOCATION : ( 13 ) . . ( 13 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES 222 > LOCATION : ( 14 ) . . ( 14 ) 223 > OTHER INFORMATION : bAla < 400 > SEQUENCE : 65 Arg Tyr Arg Arg Tyr Arg Arg Tyr Arg Arg Tyr Arg Xaa Xaa 10

< 210 > SEQ ID NO 66 < 211 > LENGTH : 14 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 13 ) . . ( 13 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _RES 22 > LOCATION : ( 14 ) . . ( 14 ) 23 > OTHER INFORMATION : bAla < 400 > SEQUENCE : 66 Arg Gly Arg Arg Gly Arg Arg Gly Arg Arg Gly Arg Xaa Xaa 10

< 210 > SEQ ID NO 67 < 211 > LENGTH : 12 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence < 220 > FEATURE : 1 > NAME / KEY : MOD _ RES < 222 > LOCATION : ( 11 ) . . ( 11 ) < 223 > OTHER INFORMATION : Acp < 220 > FEATURE : < 221 > NAME / KEY : MOD _ RES 22 > LOCATION : ( 12 ) . . ( 12 ) < 223 > OTHER INFORMATION : bAla < 400 > SEQUENCE : 67 Arg Phe Phe Arg Phe Phe Arg Phe Phe Arg Xaa Xaa 10

< 210 > SEQ ID NO 68 < 211 > LENGTH : 11 < 212 > TYPE : PRT < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : CPP sequence US 10 ,391 , 098 B2 105 106 - continued < 400 > SEQUENCE : 68 Arg Phe Phe Arg Phe Phe Arg Phe Phe Arg Gly 10

< 210 > SEQ ID NO 69 < 211 > LENGTH : 60 < 212 > TYPE : DNA < 213 > ORGANISM : Escherichia coli < 400 > SEQUENCE : 69 aaagcgagtt ttgataggaa atttaagagt atgagcacta togaagaacg cgttaagaaa 60

< 210 > SEQ ID NO 70 < 211 > LENGTH : 90 < 212 > TYPE : DNA 213 > ORGANISM : Acinetobacter baumannii < 400 > SEQUENCE : 70 ttttaa aa at ttttatattc aattaaacta gtggcaaatc a aacgccaca agcaatgagg agaattcctg tgagcgatat cgaacaacgc

< 210 > SEQ ID NO 71 < 211 > LENGTH : 11 < 212 > TYPE : DNA < 213 > ORGANISM : artificial < 220 > FEATURE : < 223 > OTHER INFORMATION : scramble targeting sequence < 400 > SEQUENCE : 71 tctcagatggt 11

35 The invention claimed is : where the targeting sequence is set forth in SEQ ID NOS : 1 . An antisense morpholino oligomer of formula ( I ) : 2 - 11 , comprises a fragment of at least 10 contiguous nucleotides of SEQ ID NOS: 2 - 11, or comprises a variant having at least 80 % sequence identity to SEQ (I ) 40 ID NOS: 2 -11 , where thymine bases ( T ) are optionally uracil bases (U ) ; X is an integer from 9 to 38 ; - Nu T is selected from OH and a moiety of the formula :

45 RO O = P - R 0 = — N (R $ )2R5 , AO ?? Lo Nu 50

where each R4 is independently C , -C , alkyl, and R is selected from an electron pair and H , and Rois selected O = P - R 55 from OH , — N ( R ) CH2C (O )NH2 , and a moiety of the formula : Nu 60 men N - R8. R2 R3 where or a pharmaceutically acceptable salt thereof, 65 R ’ is selected from H and C -Co alkyl; and where each Nu is a nucleobase which taken together R® is selected from G , C (O )R 'OH , acyl, trityl , and forms a targeting sequence , 4 -methoxytrityl , where: US 10 , 391, 098 B2 107 108 Rº is of the formula ( O - alkyl) y where y is an -continued integer from 3 to 10 and each of the y alkyl groups is independently selected from C2 - C6 alkyl; each of R ' is = N ( R !" ) , R ! ! where each R10 is inde - 5 pendently C , - Co alkyl, and Rll is selected from an ] _ electron pair and H ; N R2; R2 is selected from H , G , acyl , trityl, 4 -methoxytri tyl, benzoyl, stearoyl , and a moiety of the formula : 10 NH NI num

15 - HN NH? HN NH

HO 20 NN + R8; and (R12 N N N (R12 )2 , 25 where L is selected from C ( O )( CH2 ) . C (O ) - and C ( O )( CH2 ) 2S2 (CH2 ) 2C ( 0 ) , and each R12 is NH NH of the formula ( CH2) 20C ( O ) N (R14 ) 2where each 30 R14 is of the formula - (CH2 ) 6NHC ( NH )NH2 ; HN ? NH2 HNNH and R² is selected from an electron pair, H , and C . -C . alkyl , 35 where G is a cell penetrating peptide ( “ CPP ” ) and linker moiety selected from C ( O ) ( CH2) hd NH 5NH- CPP, C( O) ( CH2) NH- CPP, C( O ) (CH2 ) 2NHC ( O ) (CH2 ) $ NH -CPP , and C ( O ) 40 HUN CNH CH NH - CPP, or G is of the formula : ERS, 45

50 where the CPP is attached to the linker moiety by an amide bond at the CPP carboxy terminus, and NH wherein the CPP is selected from : HN . 55 NH2

HN where Ra is selected from H , acetyl, benzoyl, and stearoyl, with the proviso that R² or R $ is G , N 60 where the targeting sequence specifically hybridizes to a bacterial mRNA target sequence that encodes a protein selected from at least one of a protein associated with NH HN a biochemical pathway and /or cellular process and 165 antibiotic resistance. HNNH HNCNHI . . 2 . The antisense morpholino oligomer of claim 1 , where T is selected from : US 10 , 391 ,098 B2 109 110 109 R is of the formula : HotHO ONH2 R 'OH , and R2 is G . O = P - N ( CH3) 2 ; O = P - N (CH3 ) 2 ; 10 6 The antisense morpholino oligomer of claim 1, where T is of the formula :

= 0 1515 HOHotty

OH Z O = P - N (CH3 ) 2 , w ; and O = P - N ( CH3) 2.

and R2 is G . 7 . The antisense morpholino oligomer of claim 1 , where 3. The antisense morpholino oligomer of claim 2, where 30 T is of the formula : R2 is selected from H , G , acyl, trityl, 4- methoxytrityl , benzoyl, and stearoyl. 4 . The antisense morpholino oligomer of claim 3 , where T is selected from : 35 hotoo O = P - N ( CH3) 2. O NH2 40 min O = P — N ( CH3) 2 ; O = P - N (CH3 ) 2 ; and14 45 8 . The antisense morpholino oligomer of claim 7 , where R ? is selected from H , acyl, trityl, 4 -methoxytrityl , benzoyl, and stearoyl. 9 . The antisense morpholino oligomer of claim 8 , where ?? at least one instance of R ' is — N (CH3 ) 2 . 50 10 . The antisense morpholino oligomer of claim 9 , where each R is — N (CH3 ) 2 . 11 . The antisense morpholino oligomer of claim 1 , where and the CPP is selected from : and R² is G . 55 5 . The antisense morpholino oligomer of claim 1 , where T is of the formula : Ra . N

O = P - N ( CH3) 2. NH HN 65 | HN CNH, H?N NH ] US 10 , 391, 098 B2 111 112 -continued - continued and

HN ANH

NH NH

HN IN HNCNHNH2, HN NH L 15 NH NA

NH 20 HN NH2 NH where Ra is selected from H , acetyl, benzoyl, and 25 stearoyl. NH NH

HN NH2 HN SNH 12. The antisense morpholino oligomer of claim 1 , where G is selected from :

LL HN m A Rai

NH HN

HN CNH, H?N SNH

Z Ra; N

NH ?? womentHNCNH, H?NGNH US 10 ,391 ,098 B2 113 114 - continued HO .

N N - R " ; and S

NH * ??

HNE NH2 HN SNH

H N NH

N FR , N

HN NH2

where Rº is selected from H , acetyl, benzoyl , and stearoyl. 65 13 . The antisense morpholino oligomer of claim 1 , where the antisense oligomer is of the formula ( VII ) selected from : US 10 ,391 ,098 B2 115 116

(VIIA) VIIB)( ? ZE

IZ NH ????“? Nu 20

HN ????? (H3C)2N Nu

N(CH3)2

Nu vo

N(OH) Nu

H?N 'T

S NN HN paratofimefensagree NH 07H?N HINH

N HN HANHNH

REN. 1 ? US 10, 391 , 098 B2 117 118

(VIIG) (VIIH) -R; NSNHT. NH

N ) NH HNTNH, 07 N

(MI,C2N Nu

> N(CH3)2

N(CH3)2

-continued Nu

(H3C)2N ??????????? Nu

N(CH3)2 --S-???…???/

N Nu

N(CH3)2O NH HN. H?N

IZ HNNH, HN.

HO R US 10 ,391 ,098 B2 119 120

(VIII) Ra; (VIIJ) H2NNH. *?? -Rb; N 3' Nu 1

] NH HN?NH HN

HO. (H3C) Nu

(CH3)2|

Nu ZI NICHN(CH3)2O

.N-

-continued 3' Nu

(H3C)2N Nil Nu

N(CH3)2

anteriorment OH14 Nu

N(CH3)2 HNNH NH

NH HNANH, HN. into ?Ra- US 10 ,391 ,098 B2 121 122

(VIIK) and (VIIL)

NH2 NH HNCNH O NH Nu H? HN?

?L Last (H3C)2N Nu

CH3)2(N

N(CH32o1) Nu

-continued ? Xr

(H3C)2N Nu winbetweapopoHN N(CH3)2

N(CH3)2o

wo CZ NH2 NH NH H2NNH HN

Ro+? ??. US 10 , 391 , 098 B2 123 124 or a pharmaceutically acceptable salt of any of the fore - T is selected from OH and a moiety of the formula : going , where Ra is selected from H , acetyl, benzoyl, and stearoyl, R ' is selected from H , acetyl , benzoyl, stearoyl , trityl, and 4 -methoxytrityl , and X and Nu are 5 as defined in claim 1. 0 = ” — N ( R4) 2R5, 14 . The antisense morpholino oligomer of claim 13 , no where Rº is acetyl and R is H . 15 . The antisense morpholino oligomer of claim 1, where 10 the targeting sequence is selected from : where each R * is independently C . - C . alkyl , and R is b ) SEQ ID NO : 2 ( ATATCGCTCAC ) where X is 9 ; selected from an electron pair and H , and Rº is selected c ) SEQ ID NO : 3 (ATTCTCCTCAT ) where X is 9 ; from OH , — N ( R ) CH _ C ( O ) NH2, and a moiety of the formula : d ) SEQ ID NO : 4 (CACAGGAATTC ) where X is 9 ; 15o e ) SEQ ID NO : 5 ( TTGCCATTAGC ) where X is 9 ; f ) SEQ ID NO : 6 (CTGTAGTGATTTCACCA ) where X

m is 15 ; in N - R , g ) SEQ ID NO : 7 ( TTATCTACCAT) where X is 9 ; h ) SEQ ID NO : 8 (GCACGTTTCAT ) where X is 9 ; 20 i ) SEQ ID NO : 9 (AGAAAACCCAT ) where X is 9 ; where j ) SEQ ID NO : 10 ( TTGATAGTCAT) where X is 9 ; and R ? is selected from H and C , -C alkyl; and R8 is selected from G , - C ( O ) R ' OH , acyl, trityl, and k ) SEQ ID NO : 11 (GCTTTTTTCAT ) where X is 9 , 2538 4 -methoxytrityl , where : where thymine bases (T ) may be uracil bases ( U ) . Rº is of the formula ( O - alkyl ) y where y is an integer from 3 to 20 and each of the y alkyl groups 16 . A pharmaceutical composition , wherein the antisense is independently selected from C , - Co alkyl, morpholino oligomer is of formula ( I ) : each of Rl is — N (R192R11 where each R1° is inde pendently C , -Co alkyl, and 30 Rilis selected from an electron pair and H ; R is selected from H , G , acyl , trityl , 4 -methoxytrityl , benzoyl, stearoyl, and a moiety of the formula : Nu

O = P - R

N ' N O = P - R (R12 )2N ' N N (R12 )2 , O where L is selected from C (O ) (CH2 ) 6C ( 0 ) and La Nu 50 C ( O ) (CH2 ) 2S2( CH2 ) 2C ( O ) - , and each R12 is of the formula ( CH2 ) 2OC ( O ) N (R14 ) ,where each R14 is of the formula — ( CH2) NHC ( NH )NH2 ; and R ’ is selected from an electron pair, H , and C .- C . alkyl, R2 R3 where G is a cell penetrating peptide (“ CPP ” ) and 55 linker moiety selected from C ( O ) ( CH2) 5NH -CPP , - C (O ) (CH2 ) 2NH -CPP , - C (O ) (CH2 ) 2NHC ( O ) or a pharmaceutically acceptable salt thereof, ( CH2) -NH -CPP , and C ( O )CH NH -CPP , or G is of where each Nu is a nucleobase which taken together the formula : forms a targeting sequence , where the targeting sequence is set forth in SEQ ID NOS : 2 - 11 , comprises a fragment of at least 10 contiguous ??? , nucleotides of SEQ ID NOS : 2 - 11, or comprises a variant having at least 80 % sequence identity to SEQ ID NOS: 2 - 11 , where thymine bases ( T ) are optionally 65 uracil bases ( U ) ; X is an integer from 9 to 38 ; US 10 , 391 ,098 B2 125 126 where the CPP is attached to the linker moiety by an -continued amide bond at the CPP carboxy terminus , and wherein the CPP is selected from : H _ | - Ra.

Ra; 11 TH 10

NH NH

NH HN 15 L HN CNH, H?N CNH HN NH2 HANNH and NH

20 HUN CNH

NR, 25

w 30

Ra; 35 HN CNH, LRN 35 where Ra is selected from H , acetyl, benzoyl, and stearoyl, with the proviso that R2 or R8 is G , where the targeting sequence specifically hybridizes to 40 a bacterial mRNA target sequence that encodes a NH NH protein selected from at least one of a protein asso ciated with a biochemical pathway and /or cellular HNHNNH ? NH2 , NHONH NHNH process and antibiotic resistance . * * * * *