Oligonucleotide-based strategies to combat respiratory viruses

Abstract

The present invention relates to the field of oligonucleotide-based antiviral therapies. In particular, the invention discloses oligonucleotide molecules that may either be provided as RNA molecules capable of inducing RNAi-mediated immunity against human parainfluenza virus or as antisense oligonucleotides (ASOs), e.g., gapmers, targeting RNAs derived from human parainfluenza virus as well as oligonucleotide-based therapeutics utilizing said oligonucleotide molecules. In further aspects, the invention provides methods for screening and preparing an oligonucleotide-based therapeutic that can be used to treat an infection with a respiratory virus of interest. Such therapeutics can deliver the claimed oligonucleotides, e.g., via LNPs (lipid nanoparticles) or lipid anchors, or via lentiviral vectors or polymers such as, e.g., polyethylenimine (PEI vectors). Finally, the present invention provides a composition for use in treating a respiratory disease which comprises a lentiviral vector pseudotyped with a vesiculoviral envelope glycoprotein and at least one surface protein of influenza A virus and / or of respiratory syncytial virus.

Description

[0001]Medizinische^Hochschule^Hannover^ ^ 14104^P^6576^WO^ ^ September^2024^ Oligonucleotide-based^strategies^to^combat^respiratory^viruses^ The^present^invention^relates^to^the^field^of^oligonucleotide-based^antiviral^therapies.^In^par- ticular,^the^invention^discloses^oligonucleotide^molecules^that^may^either^be^provided^as^RNA^ molecules^capable^of^inducing^RNAi-mediated^immunity^against^human^parainfluenza^virus^ or^as^antisense^oligonucleotides^(ASOs),^e.g.,^gapmers,^targeting^RNAs^derived^from^human^ parainfluenza^virus^as^well^as^oligonucleotide-based^therapeutics^utilizing^said^oligonucleo- tide^molecules.^In^further^aspects,^the^invention^provides^methods^for^screening^and^prepar- ing^an^oligonucleotide-based^therapeutic^that^can^be^used^to^treat^an^infection^with^a^respir- atory^virus^of^interest.^Such^therapeutics^can^deliver^the^claimed^oligonucleotides,^e.g.,^via^ LNPs^(lipid^nanoparticles)^or^lipid^anchors,^or^via^lentiviral^vectors^or^polymers^such^as,^e.g.,^ polyethylenimine^(PEI^vectors).^Finally,^the^present^invention^provides^a^composition^for^use^ in^treating^a^respiratory^disease^which^comprises^a^lentiviral^vector^pseudotyped^with^a^ve- siculoviral^envelope^glycoprotein^and^at^least^one^surface^protein^of^influenza^A^virus^and / or^ of^respiratory^syncytial^virus.^ Viral^respiratory^diseases^are^among^the^most^common^diseases^in^the^world.^Respiratory^ viruses^are^efficiently^transmitted^from^person^to^person^and^affect^all^age^groups.^Many^res- piratory^diseases^occur^due^to^infections^with^various^cold^viruses^and^are^associated^with^ comparably^harmless^disease^progressions^in^most^people.^Nevertheless,^even^a^simple^cold^ often^forces^patients^to^stay^in^bed^for^several^days,^which^in^some^cases^can^lead^to^consid- erable^economic^losses^for^employers.^ In^contrast,^global^pandemics^resulting^from^infections^with^respiratory^viruses^have^repeat- edly^led^to^high^death^rates^ in^the^past.^For^example,^the^influenza^virus^subtype^A / H1N1,^ which^became^known^as^the^"Spanish^flu,"^is^estimated^to^have^claimed^between^20^and^50^ million^lives^worldwide^in^the^early^20th^century.^As^of^September^2023,^the^COVID-19^pan- demic,^ which^ halted^ public^ life^ all^ over^ the^ world,^ already^ claimed^ nearly^ 7^ million^ lives^ (https: / / covid19.who.int / ).^ In^addition,^ the^seasonal^ flu^poses^a^high^risk,^especially^ for^ the^ elderly^and / or^immunocompromised.^According^to^WHO^estimates,^flu^epidemics^lead^to^an^ estimated^3^to^5^million^cases^of^severe^illness^and^about^290,000^to^650,000^deaths^world- wide^ each^ year^ (https: / / www.who.int / news-room / fact-sheets / detail / influenza-(seasonal)).^ Moreover,^as^climate^change^progresses,^there^is^also^growing^concern^that^the^incidence^of^ viral^zoonotic^diseases,^i.e.,^infectious^diseases^that^are^caused^by^viruses^and^can^be^trans- mitted^from^animals^to^humans,^will^increase^in^the^future,^making^the^occurrence^of^further^ pandemics^more^likely.^ There^is^thus^a^growing^need^for^effective^therapies^against^respiratory^virus^infections.^ One^potential^strategy^to^combat^viral^infections^relies^on^the^use^of^nucleic^acid-based^ther- apeutics^that^target^viral^RNAs^via^complementary^Watson-Crick^base^pairing.^ Some^of^these^nucleic^acid-based^therapeutics^are^able^to^trigger^the^cleavage^or^degradation^ of^their^viral^target^RNAs,^whereas^others^effectively^block^viral^mRNA^processing^or^transla- tion.^One^group^of^nucleic^acid-based^therapeutics^relies^on^a^mechanism^known^as^RNA-^ interference^(RNAi).^RNAi^is^an^evolutionarily^conserved^defence^mechanism^which^is^used^ particularly^by^plants,^fungi,^nematodes^and^insects^to^fight^viral^infections.^During^RNAi-me- diated^antiviral^immunity,^short^pieces^of^single^stranded^ribonucleic^acid^(known^as^siRNA)^ bind^to^virus-derived^RNAs^and,^with^the^participation^of^several^enzyme^complexes,^block^ the^production^of^viral^proteins.^Since^the^RNAi^machinery^itself^is^also^conserved^in^humans^ where^it^mainly^serves^to^regulate^our^own^gene^expression,^scientists^have^recognized^the^ therapeutic^potential^of^RNAi^in^combating^viral^infections^in^human^patients^for^several^years.^ For^instance,^Bitko^et^al,^2004^discloses^inhibition^of^respiratory^viruses,^including^e.g.^RSV^ and^PIV,^using^nasally^administered^siRNA.^ Bourdreau^et^al,^2009^describes^a^therapeutic^RNAi^approach^ that^aims^to^reduce^the^ex- pression^of^a^gene.^For^this^purpose,^the^efficacy^and^toxicity^of^shRNA^and^miRNA^constructs^ were^compared^in^vitro^and^in^vivo.^The^resulting^siRNA^was^identical^in^both^cases.^Vector- based^delivery^of^ the^miRNA^constructs^was^also^ investigated.^Adeno-associated^viruses^ (AAV)^of^serotype^1^were^loaded^with^mRNA^and^shRNA^constructs^and^introduced^into^cells.^ WO^2006 / 062596^A2^describes^the^treatment^of^RSV^and^PIV^using^RNAi-inducing^dsRNA^ and^mentions^other^respiratory^viral^infections^as^possible^targets.^Intranasal^and^parenteral^ administration^of^ the^RNAi^triggers^ is^described^as^the^route^of^administration.^ In^contrast,^ delivery^of^the^active^compounds^into^the^infected^target^cells^by^viral^vector^systems^is^not^ disclosed.^ International^ patent^ application^WO^ 2006 / 110688^A9^ describes^RNAi-inducing^ constructs^ (siRNA^and^shRNA)^for^the^treatment^of^respiratory^diseases^and^explicitly^names^the^viruses^ HPIV1,^HPIV2,^HPIV3,^HPIV4a^and^HPIV4b^and^as^RNAi^targets^the^nucleotide^sequences^ of^the^(+)-^and^(-)-strand^of^the^viral^replication^cycle.^Over^8000^potentially^applicable^indi- vidual^sequences^are^described.^Also^mentioned^is^the^lentiviral^insertion^of^the^RNAi^triggers^ in^the^embodiments.^The^use^of^a^miRNA-embedded^shRNA^construct^is,^however,^not^de- scribed^in^this^publication.^ Antisense^oligonucleotides^ (ASOs)^ form^another^group^of^ nucleic^acid-based^ therapeutics^ that^are^used^in^so-called^antisense^therapy.^ASOs^are^short^single^stranded^oligonucleotides^ typically^made^of^DNA^that^are^capable^of^binding^to^a^complementary^region^within^a^target^ RNA^to^either^initiate^its^enzymatic^degradation^or^to^sterically^block^the^binding^of^cellular^ factors^or^machineries^to^the^RNA.^ 2^ ^ The^first^ASO-based^drug^to^ever^receive^FDA^approval,^Fomivirsen,^has^been^developed^for^ treating^retinitis^caused^by^cytomegalovirus^(CMV)^infection.^However,^in^light^of^the^COVID- 19^pandemic,^scientists^also^started^to^turn^to^the^development^of^ASO-based^therapeutics^ to^combat^infections^with^respiratory^viruses.^For^instance,^Zhu^et^al.,^2022^found^that^intrana- sal^administration^of^locked^nucleic^acid^ASOs^targeting^the^SARS-CoV-2^RNA^in^a^COVID- 19^mouse^model^potently^suppressed^viral^replication^in^the^lung^of^infected^mice.^ Given^the^existing^threat^of^emerging^respiratory^viral^variants,^there^is^a^need^for^new^screen- ing^methods^that^can^rapidly^and^reliably^develop^and^test^new^and^effective^nucleic^acid- based^therapeutics.^ This^problem^ is^solved^by^ the^present^ invention,^ in^particular^by^ the^subject^matter^of^ the^ claims.^ The^present^invention^provides^a^screening^method^for^an^oligonucleotide-based^therapeutic^ for^treating^an^infection^with^a^respiratory^virus^of^interest,^the^method^comprising:^ a)^ Identifying^at^least^one^target^sequence^in^an^RNA^derived^from^the^respiratory^virus^ of^interest;^and^ b)^ Generating^one^or^more^candidate^oligonucleotide^molecules^comprising^a^nucleic^ acid^sequence^complementary^to^the^at^least^one^target^sequence^identified^in^step^ a),^wherein^the^candidate^oligonucleotide^molecule^is^selected^from^the^group^con- sisting^of^an^RNA^molecule^capable^of^inducing^RNAi^and^an^antisense^oligonucleo- tide.^^ In^the^context^of^the^present^invention,^the^term^“respiratory^virus”^refers^to^a^virus^capable^of^ infecting^the^lung^and^causing^respiratory^tract^infections.^Respiratory^viruses^are^considered^ to^represent^the^most^frequent^causative^agents^of^disease^in^humans^(Boncristiani,^2009).^ The^most^common^representatives^of^ these^viruses^are^well^adapted^for^person-to-person^ transmission^and^circulate^in^a^global^scale^(Boncristiani,^2009).^ The^respiratory^virus^may^be^any^respiratory^virus^known^to^the^skilled^person,^including,^e.g.,^ influenza^virus,^respiratory^syncytial^virus,^parainfluenza^virus,^metapneumovirus,^rhinovirus,^ coronavirus,^adenovirus^and^bocavirus.^The^meaning^of^the^term^“respiratory^virus“^also^ex- tends^to^all^mutant^variants^and^serotypes^of^the^above-mentioned^viruses.^For^example,^an^ influenza^virus^may^be^an^influenza^A^virus,^e.g.,^of^strains^H5N1^or^H1N1.^For^example,^a^ coronavirus^may^be^a^SARS^virus,^e.g.,^SARS-CoV2.^ The^meaning^of^the^term^“respiratory^virus”^may^however^also^extend^to^viruses^that^are^not^ primarily^known^as^a^causing^agent^for^respiratory^tract^infections.^For^instance,^Ebola^virus^ has^been^ found^ to^ attack^ lung^ tissue^ suggesting^a^ pulmonary^ involvement^ in^Ebola^Virus^ Disease^(Lalle^et^al.,^2019).^Likewise,^Nipah^virus^has^been^shown^to^cause^person-to-person^ 3^ ^ transmittable^respiratory^infections^and^will^thus^be^considered^as^a^respiratory^virus^(Devnath^ and^Masud,^2021).^Accordingly,^a^respiratory^virus^in^the^context^of^the^invention^may^also^ encompass^Ebola^virus^and^Nipah^virus.^ In^the^context^of^the^invention,^“treating^an^infection”^is^to^be^understood^to^comprise^a^cura- tive^medical^therapy^of^a^subject^with^the^intent^to^cure,^ameliorate^or^stabilize^an^infection.^It^ is^used^synonymously^with^“treating^a^subject^having^an^infection”.^The^term^“infection”^refers^ to^the^invasion,^growth^and^proliferation^of^germs,^e.g.,^viruses,^in^the^body^of^a^subject,^which^ may^lead^to^disease.^In^some^embodiments,^treating^an^infection^is^intended^to^mean^that^the^ progression^of^ the^infection^is^ to^be^slowed,^stopped^or,^preferably,^reversed^to^allow^for^a^ perceivable^improvement^of^the^infected^subject’s^well-being^and^health.^As^used^herein,^the^ term^ “treating”^may^also^encompass^ preventing,^ i.e.,^ precluding,^ averting,^ obviating,^ fore- stalling,^stopping,^or^hindering^said^ infection^from^happening^in^the^first^place.^Treating^an^ infection^may^thus^also^explicitly^include^the^prophylactic^treatment^of^a^subject,^e.g.,^a^sub- ject^known^to^be^exposed^ to^the^virus^or^at^ risk^of^being^exposed^ to^ the^virus.^Preferably,^ treatment^is^curative.^ The^subject^of^the^herein^disclosed^invention^can^be^a^vertebrate,^such^as^a^mammal,^a^fish,^ a^bird,^a^reptile,^or^an^amphibian.^Preferably,^the^subject^of^the^herein^disclosed^methods^is^ a^mammal,^e.g.,^a^human,^ non-human^primate,^horse,^pig,^ rabbit,^dog,^sheep,^goat,^ cow,^ camel,^cat,^guinea^pig,^rat^or^mouse.^In^a^particularly^preferred^embodiment,^the^subject^is^a^ human.^ The^ term^“oligonucleotide”^ is^herein^understood^ to^ refer^ to^a^short^ polymeric^ sequence^of^ RNA^or^DNA^nucleotides.^Oligonucleotides^may^have^a^minimum^length^of^2^nucleotides^(nt)^ and^a^maximum^length^of^about^35^nt.^However,^typical^oligonucleotides^consist^of^about^12- 25^nt.^An^oligonucleotide-based^therapeutic^is^an^agent^suitable^for^pharmaceutical^use^that^ employs^oligonucleotides^capable^of^binding^with^high^specificity^to^target^RNAs^of^therapeu- tic^interest^via^complementary^Watson-Crick^base^pairing.^ The^candidate^oligonucleotide^molecules^generated^in^step^b)^may^be^an^RNA^molecule^ca- pable^of^ inducing^RNAi.^RNAi^ is^ a^natural^and^conserved^molecular^mechanism^ for^ post- transcriptional^regulation^of^gene^expression.^In^brief,^RNAi^involves^the^processing^of^dou- ble-stranded^(ds)^RNAs^into^shorter^double^stranded^fragments^of^usually^20^to^25^base^pairs^ (bp)^inside^a^cell.^Afterwards,^the^antisense^strand^of^this^dsRNA^duplex,^which^is^also^re- ferred^to^as^guide^strand,^is^incorporated^into^a^multi-protein^complex^known^as^RNA-induced^ silencing^complex^(RISC).^After^association,^this^complex^is^recruited^to^a^target^mRNA^mol- ecule^of^interest^via^complementary^base-pairing^between^the^guide^strand^and^a^target^se- quence^within^said^mRNA^molecule,^thereby^blocking^the^translation^of^the^mRNA^transcript^ into^a^polypeptide.^In^instances^where^the^antisense^strand^binds^to^its^target^sequence^with^ near^perfect^complementarity,^RNAi^promotes^the^cleavage^and^irreversible^degradation^of^ 4^ ^ the^ target^ RNA.^ Thus,^ RNAi^ can^modulate^ both^ the^ stability^ and^ translational^ efficacy^ of^ mRNAs.^^ The^RNA^molecules^capable^of^ inducing^RNAi^are^referred^herein^also^as^“RNAi-triggers”.^ RNAi^triggers^can^be^of^natural^origin^due^to^transcription^of^endogenous^genes^and^further^ processing^of^the^resulting^primary^transcripts^to^yield^the^effective^dsRNA^duplexes.^On^the^ other^hand,^RNAi^triggers^can^also^be^introduced^artificially^into^cells^via^transfection^of^RNAi^ duplexes^or^via^expression^of^RNAi^precursors^from^viral^vectors.^Since^RISC^is^specific^only^ to^RNAs^and^does^not^differentiate^between^human^and^viral^RNA,^RNAi^is^particularly^suita- ble^for^developing^therapies^against^infections^by^viruses^that^require^RNA^species^to^com- plete^their^replication^cycle.^^ In^another^embodiment^of^the^invention,^the^candidate^oligonucleotide^molecules^generated^ in^step^b)^may^be^antisense^oligonucleotides^(ASOs).^ASOs^are^artificial^single-stranded^oli- gonucleotides^of^about^12-25^nt,^more^typically^of^15-20^nt^length^that^target^RNAs^via^com- plementary^base^pairing.^They^are^usually^made^of^DNA^that^optionally^comprises^various^ chemical^modifications^to^increase^their^efficacy,^enzymatic^stability^and^decrease^their^im- munogenicity^and^off-target^ toxicity.^A^specific^ form^of^ASOs^are^so-called^gapmers^that^a^ chimeric^oligonucleotides^formed^of^a^central^short^region^made^of^DNA^nucleotides^flanked^ by^stretches^of^chemically^modified^ribonucleotides^such^as^locked^nucleic^acids^(LNAs).^ ASOs^exert^their^effects^by^two^main^mechanisms.^On^the^one^hand,^ASOs^can^hybridize^to^ their^target^RNAs^to^form^DNA-RNA^hybrids,^which^are^recognized^by^RNase^H,^an^endonu- clease^that^triggers^cleavage^of^the^RNA^strand^of^the^DNA-RNA^duplex.^Gapmers^in^partic- ular^are^specifically^designed^to^catalyze^RNase^H-dependent^degradation^of^complementary^ RNA^targets.^On^the^other^hand,^binding^of^ASOs^to^functional^cis-acting^elements^within^the^ target^RNA^may^sterically^block^access^of^the^cellular^machinery^to^the^RNA^to,^e.g.,^prevent^ translation^or^interfere^with^the^splicing^of^the^RNA.^ The^therapeutic^use^of^ASOs^is^commonly^referred^to^as^antisense^therapy.^ Because^both^RNAi-triggers^and^ASOs^are^used^to^“inhibit”,^ i.e.,^due^reduce^or^completely^ abolish^expression^of^a^target^gene^and / or^to^destroy^a^target^RNA,^the^candidate^oligonucle- otide^molecules^are^herein^also^referred^to^as^inhibitory^oligonucleotide^molecules.^ Step^a)^of^the^method^according^to^the^invention^requires^the^identification^of^sequences^in^ RNA^derived^from^a^respiratory^virus^that^are^suitable^for^being^targeted^via^RNAi^or^antisense^ therapy.^In^the^context^of^the^invention,^an^“RNA^derived^from^a^virus”^typically^is^an^mRNA^ transcribed^ from^the^genome^of^a^ respiratory^ virus.^ In^case^ the^ respiratory^virus^ does^not^ comprise^a^genome^made^of^DNA^but^rather^of^single-stranded^(ss)-^or^double-stranded^(ds)^ RNA,^the^RNA^derived^from^a^virus^may^also^be^the^genome^itself.^RNA^viruses^use^an^en- zyme^known^as^RNA-dependent^RNA^polymerase^(RdRp)^for^generating^copies^of^their^RNA^ 5^ ^ genome.^In^case^the^respiratory^virus^comprises^a^negative-sense^(-)^RNA^genome,^RdRp^is^ utilized^by^the^virus^to^produce^a^positive-sense^(+)^RNA^intermediate^which^can^be^translated^ into^protein.^Thus,^in^the^context^of^the^invention,^the^RNA^derived^from^a^virus^can^also^be^ such^a^(+)^RNA^intermediate^produced^by^negative-sense^ssRNA^viruses^for^gene^expres- sion.^ Identification^of^suitable^target^sequences^in^an^RNA^derived^from^a^virus^is^typically^achieved^ in-silico,^i.e.^by^using^a^suitable^computer^program^or^algorithm^for^automated^and^preferably^ high-throughput^alignment^of^nucleic^acid^sequences.^The^identification^of^suitable^target^se- quences^ for^RNAi^ or^ for^ antisense^ therapy^ can^be^ carried^out^ on^ the^ basis^ of^ empirically^ determined^data,^e.g.,^via^various^freely^available^online^tools,^wherein^thermodynamic^prop- erties^of^the^resulting^RNAi^or^ASO^molecule^are^taken^into^account.^Thus,^preferably,^step^ a)^of^the^screening^method^of^the^invention^is^done^in-silico^and^involves^the^use^of^an^algo- rithm^for^identifying^suitable^target^sequences^in^the^RNA^and^predicting^candidate^oligonu- cleotide^molecules^capable^of^complementary^binding^to^said^target^sequences,^wherein^the^ algorithm^takes^into^consideration^the^thermodynamic^properties^of^the^target^sequences^and^ the^predicted^candidate^oligonucleotide^molecules,^e.g.,^by^ relying^on^empirical^data^ from^ previously^ tested^oligonucleotides.^These^thermodynamic^properties^determine^or^account^ for,^e.g.,^the^formation^of^secondary^structures^in^the^target^RNA,^the^internal^stability^of^the^ oligonucleotides^complementary^to^a^target^sequence^within^said^target^RNA,^strand^asym- metry^in^case^the^candidate^oligonucleotide^molecule^is^a^dsRNA^duplex^capable^of^inducing^ RNAi,^and^the^amount^of^free^energy^of^base^pairing^between^the^target^sequence^and^the^ predicted^candidate^oligonucleotide^molecules.^^Sequence^comparison^between^different^vi- rus^strains^and^alignment^of^conserved^regions^is^usually^not^automated.^The^skilled^person^ is^aware^of^algorithms,^programs^and^online^tools^capable^of^modelling^RNA-RNA^or^DNA- RNA^interactions^to^identify^suitable^sequences^within^viral^RNA^that^can^be^targeted^using^ RNAi^or^antisense^therapy.^For^example,^the^freely^accessible^online^tools^splashRNA^(Pelos- sof^et^al.^2017)^or^the^miR30a^design^algorithm^by^Adams^et^al.^2017^can^be^used^for^this^ purpose.^^ Once^a^viral^target^RNA^sequence^is^identified^in^step^a),^one^or^more^candidate^oligonucle- otides^molecules^comprising^a^nucleic^acid^sequence^complementary^to^said^target^sequence^ are^designed^using^the^aforementioned^algorithms^and^generated^in^step^b).^^^ If^ the^candidate^oligonucleotide^molecule^is^an^RNA^molecule^capable^of^inducing^RNAi,^it^ may^be,^e.g.^a^small^interfering^RNA,^a^short^hairpin^RNA^and / or^artificial^microRNA.^ MicroRNAs^(miRNAs)^are^single-stranded,^non-coding^RNAs^that^are^ transcribed^from^en- dogenous^miRNA-genes^in^the^genome.^It^is^estimated^that^hundreds^of^distinct^and^evolu- tionary^conserved^miRNAs^modulate^the^expression^of^more^than^60%^of^all^human^protein- 6^ ^ coding^genes^within^virtually^every^biomolecular^pathway^(Catalanotto^et^al.,^2016).^Physio- logically,^miRNAs^are^generated^from^longer^endogenous^precursor^transcripts^(pri-miRNA),^ that^form^hairpin^structures^and^are^processed^through^two^distinct^endonucleolytic^cleavage^ steps^in^the^nucleus^(via^the^enzyme^Drosha)^and^cytoplasm^(via^the^enzyme^Dicer)^into^short^ RNA^duplexes^with^a^length^of^of^20-23,^e.g.,^22^base^pairs^(bp)^and^a^dinucleotide^3'^over- hang^at^both^ends.^^Based^on^its^thermodynamic^properties^defined^by^its^sequence^compo- sition,^the^antisense^strand^of^these^duplexes^is^incorporated^into^RISC^to^guide^the^complex^ to^a^complementary^target^RNA.^^Naturally^occurring^small^interfering^RNAs^(siRNAs)^rely^on^ a^similar^ effector^pathway^as^miRNAs^but^originate^ from^double-stranded^RNA^molecules^ rather^than^from^hairpin^structures.^For^ instance,^in^many^cases,^siRNAs^are^naturally^pro- duced^from^double-stranded^viral^RNA.^Mature^siRNAs^typically^have^a^length^of^20^to^25^nt^ and,^similar^to^miRNAs,^rely^on^RNAi^to^prevent^target^RNAs^from^being^translated^into^pro- tein.^ However,^whereas^ naturally^ occurring^miRNAs^ typically^ silence^ tens^ to^ hundreds^ of^ genes^by^binding^to^the^3’UTR^regions^of^the^corresponding^mRNAs^guided^by^their^8^base^ pair^“seed^region”^followed^by^repression^of^translation,^synthetic^siRNAs^typically^exhibit^a^ higher^specificity,^often^binding^to^their^target^sequences^with^up^to^100%^sequence^comple- mentarity,^ inducing^cleavage^and^degradation^of^ their^ target^mRNA^via^ the^RISC^complex^ before^translation.^Shortly^after^the^discovery^of^the^RNAi^pathway,^scientist^have^recognized^ the^ therapeutic^ potential^ of^ synthetic^ siRNA-based^ drugs.^ The^ first^ siRNA^ therapeutic,^ Patisiran,^was^approved^by^the^FDA^ in^2018^and^many^more^are^expected^to^ follow.^One^ challenge^associated^with^siRNA-based^drugs^is^site-specific^delivery.^This^requires^the^use^ of^suitable^delivery^systems^as^well^as^different^chemical^modifications^that^can^stabilize^the^ siRNA^itself.^ Short^hairpin^RNAs^(shRNA)^constitute^an^alternative^to^siRNA-based^therapeutics.^shRNAs^ are^artificially^designed^single^stranded^RNA^molecules^that^exhibit^a^characteristic^hairpin^ turn.^shRNAs^typically^have^a^length^of^50-70^nt^and^form^a^stem-loop^structure^consisting^of^ a^19^to^29^bp^region^of^double-strand^RNA^(the^stem)^bridged^by^a^region^of^predominantly^ single-stranded^RNA^(the^loop)^and^a^dinucleotide^3'^overhang.^Once^they^are^introduced^into^ a^cell,^they^are^processed^similarly^to^miRNAs^into^short^RNA^duplexes^that^resemble^siR- NAs.^One^strand^of^this^duplex^is^then^incorporated^into^RISC^to^facilitate^knock-down^of^a^ target^gene.^shRNAs^can^be^encoded^by^an^expression^vector,^such^as^a^viral^vector,^which^ allows^expression^and^processing^of^the^shRNA^construct^inside^a^target^cell.^Optionally,^they^ can^also^be^produced^by^in^vitro^transcription^of^a^linearized^plasmid-based^expression^con- struct^or^via^chemical^oligonucleotide^synthesis,^e.g.,^by^using^nucleoside^phosphoramidites^ as^building^blocks.^Additionally,^shRNAs^might^be^expressed^from^expression^plasmids,^such^ as^CpG-free^expression^vector^plasmids.^ shRNAs^are^normally^transcribed^by^RNA^polymerase^III.^However,^the^expression^of^exog- enous^shRNAs^in^cells^can^saturate^the^endogenous^miRNA^machinery,^causing^toxic^effects^ to^the^cell.^Moreover,^imprecise^processing^of^the^stem^loop^structures^of^shRNAs^may^result^ 7^ ^ in^ aberrant^ guide-^ and^ passenger-strand^mediated^ off-target^ effects.^ To^ overcome^ these^ problems,^shRNAs^may^be^provided^as^a^“shRNAmiRs”,^i.e.,^the^shRNA^is^embedded^within^ the^backbone^of^a^natural^miRNA^backbone.^Such^a^shRNAmiR^may^also^be^referred^to^as^ an^artificial^miRNA^or^miRNA^mimic.^shRNAmiRs^can^be^generated^from^Polymerase^II^pro- moters,^enabling^constitutive,^tissue^specific^or^inducible^expression^(Adams^et^al.,^2017),^as^ well^as^physiologic^expression^levels.^^ ^ Therefore,^the^candidate^oligonucleotide^molecule^produced^in^step^b)^of^the^present^inven- tion^may^also^be^an^artificial^miRNA^or^shRNAmiR.^The^skilled^person^is^aware^of^suitable^ miRNA^backbones^that^can^be^used^for^the^provision^of^shRNAs^to^cells,^which^may^include^ e.g.^the^backbone^of^miR30a,^miR223,^miR144^or^miR33.^The^shRNAmiR^constructs^may^be^ generated^as^described^in^the^example^below^or^in^Adams^et^al.,^2017,^which^is^hereby^in- corporated^by^reference,^and^accordingly,^may^have^a^design^that^corresponds^to^the^design^ of^the^shRNAmiR^constructs^disclosed^in^Adams^et^al.,^2017.^In^brief,^ the^shRNAmiR^con- structs^preferably^comprise^flanking^regions^and,^downstream^of^the^stem^loop^of^the^miRNA,^ a^binding^sequence^that^is^complementary^to^the^RNA^target^sequence^identified^in^step^a)^ of^ the^method^according^ to^ the^ invention.^A^sequence^complementary^ to^said^binding^se- quence^may^be^arranged^upstream^of^the^stem^loop^of^the^miRNA,^in^order^to^allow^formation^ of^a^hairpin^structure.^The^stem^loop^sequence^may^be^adjusted^according^to^the^respective^ miRNA^backbone^used.^Polycistronic^line^up^of^different^miRNA^backbones^in^different^com- binations^may^be^performed^to^achieve^parallel^knockdown^of^ several^ targets.^These^may^ consist^of^different^variations^of^miRNA^backbones^miR30a,^miR223,^miR33^and / or^miR144^ in^double,^triple^or^quadruple^constructs^(exemplified^in^Table^1).^In^between^the^backbones,^ spacer^regions^of^0-99^base^pairs^of^noncoding^RNA^may^be^introduced^into^the^lineup.^^ Table^1:^Exemplary^combinations^of^different^miRNA^backbones^that^may^be^combined^in^a^ single^shRNAmiR^construct.^ ^ 5 m ;^ ;^ ;^ m ;^ ;^ ;^ miR144_miR30a_miR33^ 8^ ^ miR33^ miR30a;^ miR223;^ miR144;^ miR30a_miR223;^ miR30a_miR144;^ ;^ ;^ m ;^ ;^ ;^ ^ The^candidate^oligonucleotide^molecule^generated^in^step^b)^comprises^a^nucleic^acid^se- quence^that^ is^complementary^to^a^target^sequence^identified^in^step^a)^of^ the^method^ac- cording^to^ the^ invention.^ In^ the^context^of^ the^ invention,^ this^nucleic^acid^sequence^ is^also^ referred^to^as^“guide^sequence”.^In^case^the^candidate^oligonucleotide^molecule^is^an^siRNA^ or^an^ASO,^ the^candidate^oligonucleotide^molecule^may^ also^consist^of^such^a^ guide^ se- quence.^In^the^context^of^the^invention,^a^guide^sequence^is^considered^complementary^to^its^ target^sequence^when^it^has^been^designed^to^at^least^partially^hybridize^to^said^target^se- quence.^Hybridization^refers^to^a^process^wherein^two^single-stranded^nucleic^acid^molecules^ anneal^ to^each^other^via^complementary^Watson–Crick^base^pairing.^ In^the^case^of^DNA- RNA^interactions,^the^nitrogenous^base^adenine^present^in^the^DNA^strand^thus^pairs^and^ forms^hydrogen^bonds^with^uracil^in^the^opposing^RNA^strand^strand,^whereas^adenine^pre- sent^in^the^RNA^pairs^with^thymine^in^DNA.^In^the^case^of^RNA-RNA^interactions,^the^nitrog- enous^base^adenine^present^in^one^RNA^strand^pairs^and^forms^hydrogen^bonds^with^uracil^ in^the^opposing^strand.^Regardless^of^whether^the^interactions^occur^between^DNA^and^RNA^ or^between^two^RNA^molecules.cytosine^pairs^with^guanine.^Accordingly,^the^guide^sequence^ is^complementary^to^the^target^sequence^if^it^comprises^a^sequence^of^complementary^bases^ mirroring^at^least^a^part^of^the^target^sequence,^such^that^it^allows^for^RNAi^or^ASO-mediated^ RNA^degradation / blockage.^ In^one^embodiment,^the^candidate^oligonucleotide^molecule^consists^of^or^comprises^a^guide^ sequence^that^is^100%^complementary^to^its^target^sequence,^ i.e.,^the^guide^sequence^hy- bridizes^over^its^entire^length^to^the^target^RNA^sequence,^wherein^each^base^of^the^guide^ sequence^hybridizes^to^a^corresponding^complementary^base^of^the^target^RNA^sequence.^ In^such^a^scenario,^the^candidate^oligonucleotide^molecule^is^highly^specific^to^its^target^se- quence^and^ thus^ typically^does^not^bind^ to^other^sequences^within^a^given^sample.^ If^ the^ candidate^oligonucleotide^candidate^is^an^RNA^capable^of^inducing^RNAi,^the^guide^sequence^ is^preferably^20-23^nt,^e.g.,^22^nt^long^and^fully^complementary^to^the^target^sequence.^If^the^ candidate^oligonucleotide^ ins^an^ASO,^the^guide^sequence^is^preferably^12-16^nt^and^fully^ complementary^to^the^target^sequence.^ 9^ ^ However,^in^some^embodiments,^it^may^be^preferable^that^at^least^1,^e.g.,^at^least^2,^at^least^ 3,^at^least^4,^or^at^least^5^bases^of^the^guide^sequence^have^mismatches^with^their^ target^ RNA^sequence,^ i.e.,^ the^guide^sequence^does^not^ bind^with^100%^complementarity^ to^ its^ corresponding^target^sequence.^Accordingly,^the^guide^sequence^of^the^candidate^oligonu- cleotide^molecule^may^have^a^nucleic^acid^sequence^that^is^merely^75-99%,^e.g.,^75%,^80%,^ 85%,^90%,^95%^or^99%^complementary^to^its^target^sequence.^In^such^a^scenario,^the^can- didate^ oligonucleotide^molecule^still^exhibits^a^very^high^specificity^ to^ its^ target^ sequence,^ while^also^allowing^the^recognition^of^variants^thereof.^This^can^be^advantageous^because^of^ viral^variability,^particularly^observed^after^mutations^in^the^viral^genome^or^for^members^of^ the^same^viral^family.^ In^another^embodiment,^the^guide^sequence^of^the^candidate^oligonucleotide^molecule^hy- bridizes^to^the^target^RNA^with^only^a^relatively^short^stretch^of^its^sequence.^For^example,^ naturally^occurring^miRNAs^generally^do^not^hybridize^to^a^target^RNA^over^their^entire^length.^ Rather,^miRNAs^only^bind^to^their^target^mRNA^with^a^short^stretch^of^6^to^8^nucleotides^(nt)^ at^their^5'^end^(the^so-called^seed^sequence).^Accordingly,^the^guide^sequence^of^a^candidate^ oligonucleotide^molecule^may^already^be^considered^complementary^ to^a^target^sequence^ within^the^meaning^of^the^present^invention^when^it^is^complementary^to^at^least^a^seed^region^ of^at^least^5^to^15,^e.g.,^5,^6,^7,^8,^9,^10,^11,^12,^13,^14,^or^15^nucleotides^to^its^respective^ target.^^ The^candidate^oligonucleotide^molecules^can^be^generated^by^any^suitable^method.^For^ex- ample,^the^oligonucleotide^molecules^can^be^generated^by^chemical^synthesis,^which^is^typ- ically^done^by^commercial^companies^on^a^customized^basis.^Alternative^methods^for^gener- ating^ in^particular^RNA^molecules^capable^of^ inducing^RNAi^are^well^ known^ to^ the^skilled^ person^and^ include,^e.g.,^ in^vitro-transcription^or^ in^ vivo^ transcription^ from^DNA^ templates^ introduced^into^cells.^^ In^some^embodiments,^the^candidate^oligonucleotide^molecules^generated^in^step^b)^of^the^ method^of^the^invention^can^be^directly^incorporated^into^an^oligonucleotide-based^therapeu- tic^ for^ treating^an^ infection^with^ a^ respiratory^virus^of^ interest,^as^described^ in^more^detail^ below.^However,^preferably,^the^candidate^oligonucleotide^molecules^generated^in^step^b)^of^ the^method^according^to^the^invention^are^further^screened^for^their^knockdown^efficiencies^ and / or^their^ability^to^induce^a^reduction^of^the^titer^of^^the^respiratory^virus^of^interest.^ Disclosed^herein^ but^ not^ claimed^ as^ part^ of^ the^ present^ invention^ is,^ e.g.,^ an^ “exemplary^ screening^step^c)”^after^step^b)^comprising^screening^the^candidate^oligonucleotide^molecules^ generated^in^step^b)^for^their^ability^to^induce^a^reduction^of^the^titer^of^the^respiratory^virus^of^ interest^in^lung^tissue,^wherein^the^screening^comprises^steps^of^ i.^ providing^cut^slices^obtained^from^lung^tissue,^^ 10^ ^ ii.^ Introducing^the^one^or^more^candidate^oligonucleotide^molecules^generated^in^ step^b)^into^the^cut^slices,^ iii.^ Infecting^the^cut^slices^with^the^respiratory^virus^of^interest,^ iv.^ Obtaining^supernatant^ from^ the^ cut^ slices^at^a^ first^ time^point^ and^at^ least^a^ second^time^point^after^infection;^ v.^ Determining^the^respiratory^virus^titre^in^the^supernatants^obtained^at^the^first^ time^point^and^at^the^at^least^second^time^point^of^step^ iv^for^each^candidate^ oligonucleotide^molecule,^ wherein^steps^c)^ii.^and^iii.^may^be^performed^in^any^order.^ Since^the^oligonucleotide-based^therapeutic^preferably^is^for^use^in^a^human^subject,^the^lung^ tissue^used^for^preparing^the^cut^slices^is^preferably^of^human^origin^as^well.^The^lung^tissue^ should^be^viable,^e.g.,^viable^human^lung^tissue.^The^lung^tissue^thus^preferably^is^human^ explant^lung^tissue^that^has^been^surgically^removed^from^a^human^subject^and^is^grown^in^ an^explant^culture.^ However,^many^respiratory^viruses,^including^e.g.,^influenza^A^virus^strains^H5N1^or^H1N1,^ are^known^to^infect^animals,^in^particular^farm^animals^such^as^chickens^or^pigs.^The^screen- ing^of^the^candidate^oligonucleotide^molecules^may^thus^also^be^performed^in^lung^tissues^of^ animal^ origin,^ e.g.,^ lung^ tissues^ from^pigs,^cattle,^ sheep,^mice,^ rats,^cotton^ rats,^ ferrets^or^ chickens.^ The^preparation^of^ the^ lung^cut^slices,^ referred^herein^also^as^precision-cut^ lung^slices^or^ PCLS,^ can^be^done^as^described^ in^Danov^ et^ al.,^ 2019,^which^ is^ hereby^ incorporated^by^ reference^in^its^entirety.^Briefly,^human^explant^lung^tissue^is^inflated^by^being^filled^with^aga- rose.^After^polymerization^of^the^agarose,^the^ lung^ tissue^is^punched^and^cut^with^a^tissue^ slicer^to^obtain^200^–^300^µm^thick^round^slices^with^a^diameter^of^about^8mm.^The^skilled^ person^will^be^aware^of^alternative^methods^for^producing^lung^cut^slices^of^comparable^thick- ness.^Accordingly,^the^production^of^the^lung^cut^slices^is^not^necessarily^limited^to^the^method^ as^described^to^Danov^et^al.,^2019.^ The^freshly^prepared^lung^cut^slices^are^provided^in^a^suitable^culture^medium,^e.g.,^DMEM / F- 12^supplemented^with^penicillin / streptomycin,^and^may^be^maintained^under^typical^cell^cul- ture^conditions.^e.g.,^at^a^temperature^of^37°^at^5%^CO2^for^7-10^days.^ In^step^ ii)^ the^candidate^oligonucleotide^molecules^are^ introduced^ into^ the^ lung^cut^slices,^ which^may^be^achieved^by^any^suitable^method^known^to^the^skilled^person.^For^instance,^ the^candidate^oligonucleotide^molecules^may^be^introduced^by^means^of^transfection,^e.g.,^ by^lipofection,^calcium^phosphate^transfection^or^electroporation.^^ 11^ ^ Alternatively,^ the^candidate^oligonucleotide^molecules^may^be^ introduced^ into^the^ lung^cut^ slices^using^a^ suitable^vector.^ For^ instance,^ if^ the^candidate^ oligonucleotide^ is^ one^of^ the^ herein-described^RNAi-trigger^the^vector^may^be^an^expression^vector^encoding^said^RNAi^ trigger.^Examples^of^such^expression^vectors^include^but^are^not^limited^to^minicircles,^plas- mids,^cosmids,^phages,^viruses^or^artificial^chromosomes.^In^some^embodiments,^the^vector^ preferably^is^a^viral^vector,^e.g.,^a^lentiviral^vector,^an^adenoviral^vector^or^an^adeno-associ- ated^viral^vector.^Expression^vectors^typically^contain^a^cargo^sequence^comprising^an^ex- pression^cassette,^i.e.^the^necessary^elements^that^permit^transcription^of^a^nucleic^acid^into^ the^candidate^RNA^molecule,^such^as^a^suitable^promoter.^ The^ ideal^means^ for^ introducing^ the^candidate^ oligonucleotide^molecule^ into^ the^ lung^ cut^ slices^largely^depends^on^whether^the^candidate^oligonucleotide^molecules^are^provided^as^ siRNAs^and^ASOs^or^shRNAs / shRNAmiRs.^In^case^the^candidate^oligonucleotide^molecules^ are^siRNAs^or^ASOs,^they^usually^do^not^require^further^endonucleolytic^processing^to^exert^ their^biological^effect.^Accordingly,^they^may^be^directly^delivered^into^the^lung^cut^slices^via^ transfection,^preferably^via^lipofection.^^ On^the^other^hand,^candidate^oligonucleotide^molecules^that^are^shRNAs^or^shRNAmiRs^are^ preferably^introduced^into^the^lung^cut^slices^using^a^suitable^expression^vector,^preferably^a^ viral^vector.^They^may^however^also^be^introduced^by^different^means,^e.g.,^via^transfection,^ e.g.,^via^lipofection^of^in^vitro-transcribed^shRNAmiR^constructs.^ After^the^introduction^of^the^candidate^oligonucleotide^molecules,^the^lung^cut^slices^are^kept^ at^a^temperature^of^32^to^40°C,^e.g.,^32,^33,^34,^35,^36,^37,^38,^39^or^40°C,^preferably^at^35°C^ before^being^infected^with^a^suitable^amount^of^the^respiratory^virus^of^interest,^according^to^ standard^ infection^protocols.^For^instance,^ the^ lung^cut^slices^may^be^infected^with^a^virus^ titre^of^ ^2*102^ -2*106,^e.g.,^ of^2*102-^2*105,^ 2*103-2*106,^ 2*102-^2*104,^2*103-^2*105,^2*104-^ 2*106^or^of^2*102,^2*103,^2*104,^2*105^or^2*106^focus^forming^units^(FFU)^in^200µL.^Infection^of^ the^ lung^cut^slices^with^ the^ respiratory^ virus^of^ interest^may^ take^place,^ e.g.,^about^ 12-96^ hours,^e.g.^about^12,^24,^36,^48,^60,^72,^84^or^96^hours^after^introduction^of^the^candidate^ oligonucleotide^molecules.^Preferably,^ the^ lung^cut^slices^are^ infected^with^ the^ respiratory^ virus^of^ interest^24-84^hours^after^ introduction^of^ the^candidate^oligonucleotide^molecules.^ The^time^between^the^introduction^of^the^candidate^oligonucleotide^molecules^and^viral^infec- tion^may^depend^on^the^means^for^introducing^the^candidate^oligonucleotide^molecule^into^ the^ lung^ cut^ slices.^ For^ instance,^ if^ the^ candidate^ oligonucleotide^ molecules,^ e.g.^ shRNAs / shRNAmiRs,^are^transduced^with^a^viral^vector^into^the^lung^cut^slices,^infection^with^ the^respiratory^virus^of^interest^may^preferably^be^done^about^72^hours^afterwards^to^provide^ sufficient^time^for^the^expression^and^endonucleolytic^processing^of^the^candidate^oligonucle- otides^molecules,^e.g.^shRNA / shRNAmiR,^ inside^ the^cut^slices.^If,^however,^ the^candidate^ oligonucleotide^molecules^are^ transfected,^e.g.,^as^siRNAs^or^ASOs^into^the^cell,^ infection^ with^the^respiratory^virus^of^interest^may^be^done^earlier,^e.g.^already^about^24^hours^after^ 12^ ^ introduction^of^the^candidate^oligonucleotide^molecule.^The^skilled^person^will^have^no^prob- lem^to^determine^the^ideal^time^point^for^infecting^the^cell^with^the^respiratory^virus^of^interest^ within^the^limits^of^the^present^specification.^It^may^be^even^advantageous^to^introduce^the^ candidate^oligonucleotide^candidates^into^the^lung^cut^slices^while^simultaneously^infecting^ them^with^the^respiratory^virus.^ It^ is^also^possible^to^ first^ infect^ the^ lung^cut^slices^with^the^ respiratory^virus^of^ interest^before^ introducing^the^candidate^oligonucleotide^molecule.^For^ instance,^the^candidate^oligonucleotide^molecules^can^be^introduced^into^the^lung^cut^slices^ about^12-36^hours,^e.g.,^24^hours,^after^they^were^infected^with^the^respiratory^virus^of^inter- est.^After^introducing^the^candidate^oligonucleotide^molecule^and^infecting^the^lung^cut^slices^with^ the^respiratory^virus^of^interest,^culture^supernatant^is^obtained^from^the^cultured^cut^slices^at^ a^first^time^point^and^at^least^a^second^time^point.^ The^first^time^point^should^ideally^be^chosen^shortly^after^infecting^the^lung^cut^slices^with^the^ respiratory^virus^of^interest.^For^instance,^the^first^time^point^may^be^30^minutes^to^2^hours^ after^infection.^Preferably,^the^supernatant^is^obtained^from^the^cut^slices^at^a^first^time^point^ of^about^1^hour^after^infection.^In^case^the^lung^slices^were^first^infected^with^the^virus^prior^to^ introduction^of^the^candidate^oligonucleotide^molecules,^the^first^time^point^may^be^before^or^ after^introduction^of^the^candidate^oligonucleotide^molecules,^preferably,^shortly^before^said^ introduction,^e.g.,^about^one^hour^before^said^introduction.^ The^second^time^point^should^ideally^be^chosen^to^be^about^24-96^hours,^e.g.^about^12,^24,^ 36,^48,^60,^72,^84^or^96^hours^after^the^first^time^point,^and^in^any^case,^at^least^6^hours^after^ introduction^of^the^candidate^oligonucleotide^molecules.^Preferably,^the^second^time^point^is^ about^48^to^72^hours,^e.g.,^about^48,^60^or^72^hours^after^the^first^time^point.^ Afterwards,^the^respiratory^virus^titre^in^the^supernatants^obtained^at^the^first^time^point^and^ at^the^at^least^second^time^point^is^determined^for^each^candidate^oligonucleotide^molecule^ to^identify^preferred^candidates^that^most^effectively^reduced^viral^load^between^the^two^time^ points.^In^the^context^of^the^invention,^viral^load^is^considered^to^be^“effectively^reduced”^in^ cases^where^the^virus^titre^in^the^supernatant^obtained^at^the^second^time^point^ is^reduced^ by^at^least^25%,^preferably,^by^at^least^35%,^more^preferably^by^at^least^50%^compared^to^ the^virus^titre^ in^the^supernatant^obtained^at^ the^first^ time^point.^For^ instance,^viral^ load^is^ considered^to^be^“effectively^reduced”,^if^the^viral^titre^in^the^supernatant^of^the^second^time^ point^is^reduced^by^more^than^50%^compared^to^the^viral^titre^in^the^supernatant^of^the^first^ time^point,^e.g.,^by^at^least^60,^70,^80,^85,^90,^95^or^99%.It^may^be^particularly^preferred^if^the^ virus^titre^ in^the^supernatant^obtained^at^the^second^time^point^has^been^reduced^by^100%^ compared^to^the^virus^titre^of^the^first^supernatant,^i.e.,^if^no^traces^of^respiratory^virus^can^be^ detected^at^all^in^the^supernatant^obtained^at^the^second^time^point.^^ 13^ ^ Determining^the^virus^titre^may^be^done^by^any^suitable^method^known^to^the^skilled^person,^ e.g.,^ it^may^be^determined^using^a^focus^forming^assay^(FFA)^ in^cell^culture.^Supernatants^ may^also^be^collected^from^ the^cultured^ lung^cut^slices^at^additional^time^points,^e.g.,^at^a^ third,^fourth^or^even^fifth^timepoint^to^monitor^the^effect^of^the^candidate^oligonucleotide^mol- ecules^on^viral^ titre^over^a^longer^period^of^ time.^The^candidate^oligonucleotide^molecules^ that^exhibited^the^most^effective^reduction^in^viral^load^during^the^above^described^exemplary^ step^c)^are^then^preferably^selected^for^producing^the^oligonucleotide-based^therapeutic.^ As^already^indicated,^the^above-mentioned^“exemplary^screening^step^c)”^is^not^part^of^the^ present^method^of^the^invention^but^has^only^been^described^for^illustrative^purposes.^ In^other^words,^the^candidate^oligonucleotide^molecules^generated^in^step^b)^of^the^method^ of^ the^ invention^are^not^subjected^ to^the^above-described^exemplary^screening^step^c).^ In^ some^embodiments,^they^may^however^be^subjected^to^one^or^more^alternative^screening^ steps.^^ Such^an^alternative^screening^step^c)^may,^in^some^embodiments,^comprise:^ i.^ Cloning^an^RNA^sequence^ identified^ in^step^a)^ or^parts^ thereof^ into^a^UTR^of^an^ mRNA^molecule^encoding^a^fluorescent^protein^to^obtain^a^reporter^mRNA;^ ii.^ Introducing^said^reporter^mRNA^into^a^cell,^ iii.^ Introducing^the^one^or^more^candidate^oligonucleotide^molecules^generated^in^step^ b)^into^the^cell,^and^ iv.^ Assessing^the^fluorescence^intensity^ in^the^cells^comprising^the^reporter^mRNA^to^ determine^a^knock-down^efficiency^for^each^candidate^oligonucleotide^molecule.^ ^ In^the^context^of^the^present^invention,^this^alternative^screening^step^c)^relying^on^a^reporter^ mRNA^is^also^referred^to^as^“first^screening^step”.^ Fluorochrome-based^knock-down^(KD)^reporter^assays^used^for^validating^RNAi^efficiency^ and^potency^of^synthetic^RNAi-triggers^are^known^from^the^state^ in^the^art^ (e.g.,^Du^et^al.,^ 2004;^Kumar^et^al.,^2003,^Fellmann^et^al.,^2011).^Of^note,^these^assays^may^also^be^used^to^ assess^the^knock-down^efficiency^of^ASOs.^Briefly,^an^RNA^target^sequence^identified^in^step^ a)^of^the^method^of^the^invention^or^a^part^thereof^is^placed^into^an^untranslated^region^(UTR)^ of^a^typically^constitutively^expressed^mRNA^encoding^a^fluorescent^protein,^such^as,^e.g.,^ BFP,^GFP,^mCherry^or^Venus.^Because^the^target^sequences^of^miRNAs^are^usually^located^ within^the^3’UTR^of^an^mRNA,^the^target^sequence^identified^in^step^a)^ is^preferably^ intro- duced^into^the^3’UTR^of^the^reporter^mRNA^as^well.^^ The^reporter^mRNA^is^subsequently^ introduced^in^step^ii)^of^the^”first^screening^step”^into^a^ cell^in^culture^such^as,^e.g.,^a^32D^cell^or^a^293T^cell.^For^instance,^the^reporter^mRNA^may^ 14^ ^ be^an^ in^ vitro^ transcribed^mRNA^ that^ is^ introduced^ into^ the^cell^by^ a^suitable^ transfection^ method^such^as^lipofection.^Alternatively,^a^suitable^expression^vector,^as^described^herein,^ may^be^used^for^introducing^a^transgene^that^expresses^the^reporter^mRNA^inside^the^cell^ under^the^control^of^a^suitable^promoter.^Said^promoter^may^be^configured^to^facilitate^con- stitutive^ transcription^of^ the^ reporter^mRNA^within^ the^cell,^ resulting^ in^varying^amounts^of^ reporter^transcripts^depending^on^the^strength^of^the^promoter.^This^facilitates^adjustable^sen- sitivity^towards^the^tested^constructs.^For^instance,^the^promoter^may^be,^e.g.,^an^SFFV^or^an^ EFS^promoter.^The^promoter^may^alternatively^also^be^an^inducible^promoter,^such^as^a^tet- racycline-inducible^promoter^that^triggers^transcription^of^the^reporter^mRNA^only^in^the^pres- ence^of^an^external^stimulus,^such^as^the^addition^of^tetracycline^or^doxycycline.^The^skilled^ person^will^have^no^problems^ to^select^suitable^expression^vectors^and^promoters^ for^ex- pressing^the^reporter^mRNA^inside^a^cell.^The^vector^may^also^be^configured^to^integrate^the^ transgene^into^the^genome^of^the^cell^to^obtain^a^reporter^cell^line^that^stably^expresses^the^ reporter^mRNA.^ For^the^purpose^of^the^present^invention,^the^above-described^reporter^mRNA^assay^is^pref- erably^further^optimized^as^described^in^Adams^et^al.,^2017.^For^this,^the^UTR,^e.g.^the^3’UTR^ of^ the^ reporter^mRNA^does^not^only^comprise^ a^single^cognate^RNA^ target^sequence^but^ rather^a^plurality^or^even^all^of^the^target^sequences^identified^in^step^a)^of^the^method^ac- cording^to^the^invention.^As^described^in^Adams^et^al.^2017,^the^plurality^of^RNA^target^se- quences^can^be^incorporated^en^bloc^into^the^3'UTR^of^the^reporter^mRNA,^e.g.,^by^using^the^ gBlocks™-technology^provided^by^Integrated^DNA^Technologies^(IDT).^This^way,^the^authors^ of^Adams^et^al.,^2017^managed^to^concatenate^up^to^65^shRNA^target^sequences^in^the^3’^ UTR^of^a^BFP^fluorescence^reporter^gene^(designated^as^mTagBFP2).^By^including^multiple^ target^sites^within^a^single^UTR^of^a^reporter^mRNA,^it^is^not^necessary^to^generate^a^separate^ reporter^mRNA^construct^and / or^reporter^cell^ line^for^each^candidate^oligonucleotide^mole- cule.^Rather,^cells^of^a^single^reporter^cell^line^comprising^and / or^expressing^the^herein^de- scribed^reporter^mRNA^may^be^used^to^screen^a^plurality^or^even^all^of^the^candidate^oligo- nucleotide^molecules^generated^ in^step^b)^of^ the^method^according^to^the^ invention.^ ^This^ allows^for^time-saving^parallel^candidate^screening,^as^each^candidate^oligonucleotide^mole- cule^to^be^screened^can^be^simply^introduced^into^a^separate^cell^of^the^same^reporter^cell^ line.^Additionally,^the^reporter^mRNA^construct^may^be^equipped^with^an^antibiotic-resistance^ cassette^such^as,^e.g.,^a^puromycin^resistance^cassette,^to^enable^positive^selection^of^re- porter-positive^cells^prior^to^the^validation^process.^In^case^the^reporter^construct^comprises^ a^puromycin^resistance^cassete,^selection^may^take^place^by^supplementation^with^about^0.4^ –^4^µg / mL^puromycin^which^typically^killes^at^least^90%^of^reporter-negative^cells.^ The^candidate^oligonucleotide^molecule(s)^generated^in^step^b)^of^the^method^according^to^ the^invention^is^next^introduced^into^the^cell^as^already^described^herein,^wherein^each^can- didate^oligonucleotide^molecule^is^introduced^into^a^separate^cell^comprising^and / or^express- ing^the^reporter^mRNA.^Thus,^if^the^candidate^oligonucleotide^molecule^is^an^siRNA^or^ASO,^ 15^ ^ it^is^preferably^introduced^via^transfection,^e.g.,^via^lipofection.^For^instance,^the^siRNA / ASO^ may^be^introduced^into^the^cell^using^lipid^nanoparticles^(LNPs).^Further^methods^for^intro- ducing^siRNA^molecules^or^ASOs^into^a^target^cell^are^known^to^the^skilled^person.^For^ex- ample,^siRNA^molecules^or^ASOs^may^also^be^introduced^using^cationic^polymers^such^as,^ e.g.,^polyethylenimine^(PEI)^or^DEAE-dextran,^or^branched^polymeric^molecules^known^as^ dendrimers.^The^ siRNA^or^ASO^may^alternatively^also^directly^be^ labelled^with^ a^ suitable^ lipophilic^anchor,^e.g.,^by^directly^conjugating^ the^siRNA^or^ASO^ to^ lipid^residues^such^as^ palmityl-^or^cholesterol-residues.^In^cases^where^the^candidate^oligonucleotide^molecule^is^ provided^as^an^shRNA^or^an^shRNAmiR,^it^is^preferably^introduced^by^means^of^a^viral^vector^ such^as^a^ lentiviral^vector.^ Like^siRNAs^or^ASOs,^ however,^ in^ vitro-transcribed^shRNA^or^ shRNAmiR^may^also^be^transfected^into^the^the^cells,^e.g.,^via^lipofection^using,^e.g.,^LNPs^ or^via^cationic^polymers^such^as,^e.g.^PEI.^ In^the^absence^of^an^RNAi^trigger / ASO^or^in^case^a^candidate^oligonucleotide^molecule^intro- duced^in^step^iii)^does^not^or^only^weakly^bind^to^any^of^the^RNA^target^sequences^within^the^ UTR^of^ the^ reporter^mRNA,^ the^reporter^mRNA^ is^ translated^ into^ the^encoded^ fluorescent^ protein^of^choice.^Accordingly,^a^corresponding^fluorescent^signal^can^be^detected^by^meth- ods^known^to^ those^skilled^ in^ the^art,^e.g.,^ via^ fluorescence-activated^cell^ sorting^ (FACS),^ fluorescence^microscopy^or^western^blotting.^If,^however,^a^candidate^oligonucleotide^mole- cule^that^is^to^be^screened^using^the^“first^screening^step”^comprises^a^guide^sequence^com- plementary^to^any^of^the^target^RNA^sequences^in^the^UTR^of^the^reporter^mRNA,^it^is^ex- pected^to^prevent^its^translation^into^a^fluorescent^protein^by^triggering^its^RNAi-^or^antisense- mediated^knock-down.^In^consequence,^the^fluorescent^signal^produced^by^the^reporter^cell^ will^be^reduced^or^completely^abolished^(i.e.^the^signal^will^be^below^the^detection^limit^of^the^ method^used^for^measuring^the^fluorescent^signal).^By^assessing^the^fluorescence^intensity^ in^the^cells^comprising^the^reporter^mRNA^that^have^been^exposed^to^a^given^candidate^oli- gonucleotide^molecule,^the^skilled^person^will^be^able^to^easily^determine^and^calculate^the^ knock-down^efficiency^for^each^candidate^oligonucleotide^molecule^based^on^standard^meth- ods.^This^can^be^achieved,^e.g.,^by^comparing^the^effect^of^a^given^candidate^oligonucleotide^ molecule^on^fluorescence^intensity^in^the^cells^comprising^the^reporter^mRNA^to^one^or^more^ reference^or^control^shRNA / ASO^constructs^of^known^knock-down^efficiency.^Suitable^refer- ence^shRNAs^have^been^reported^in^the^state^of^the^art^and^include,^e.g.,^shRNAs^against^ Pten^or^Cebpa^as^described^in^Fellmann^et^al.,^2011.^Of^course,^to^be^used^as^controls,^the^ UTR^of^the^reporter^mRNA^must,^in^addition,^also^comprise^the^target^sequence^of^said^ref- erence^shRNAs.^ Typically,^fluorescent^intensity^in^the^cells^comprising^the^reporter^mRNA^is^assessed^about^ 12-72^hours,^preferably^24-60^hours,^most^preferably^48^hours^after^introduction^of^the^can- didate^oligonucleotide^molecules^to^provide^sufficient^time^for^reporter^knock-down^and^deg- radation^of^sufficient^amounts^of^residual^fluorescent^protein^in^the^cytoplasm.^ 16^ ^ In^general,^a^strong^fluorescent^signal^and^thus^a^high^fluorescence^intensity^in^relation^to^the^ one^or^more^reference^or^control^shRNAs / ASOs^is^associated^with^a^weak^knock-down^effi- ciency^of^a^candidate^oligonucleotide^molecule,^whereas^a^weak^fluorescent^signal^and^thus^ a^low^fluorescent^intensity^in^relation^to^the^one^or^more^reference^or^control^shRNAs / ASOs^ is^associated^with^a^strong^knock-down^efficiency^of^a^candidate^oligonucleotide^molecule.^ At^the^end^of^the^herein^disclosed^“first^screening^step”,^a^candidate^oligonucleotide^molecule^ exhibiting^a^“strong”^knock-down^efficiency,^ i.e.,^an^knock-down^efficiency^of^about^at^ least^ 70%,^preferably^about^at^least^80%,^and^most^preferably^at^about^at^least^90%^as^determined^ by^the^fluorescence^intensity^in^the^cells^comprising^the^reporter^mRNA^may^be^considered^ suitable^for^use^in^an^oligonucleotide-based^therapeutic^for^treating^an^infection^with^the^res- piratory^virus^of^interest.^ The^method^of^the^invention^may^alternatively^or,^preferably,^in^addition^to^the^“first^screening^ step”^involving^the^reporter^mRNA^described^herein,^comprise^another^screening^step^after^ step^b).^In^the^context^of^the^invention,^this^additional^or^alternative^screening^step^is^herein^ also^referred^to^as^^“second^screening^step”^and^may^comprise^ I.^ Introducing^the^one^or^more^candidate^oligonucleotide^molecules^generated^in^step^b)^ into^a^cell^in^cell^culture,^ II.^ ^Infecting^the^cell^of^(I)^with^the^respiratory^virus^of^interest,^ III.^ Obtaining^supernatant^from^the^cell^of^(I)^at^a^first^time^point^and^at^least^a^second^time^ point^after^infection;^ IV.^ Determining^the^virus^titre^in^the^supernatants^obtained^at^the^first^time^point^and^at^ the^at^least^second^time^point^of^step^(III).^ The^“second^screening^step”^resembles^the^above-described^“exemplary^screening^step^c)”,^ with^ the^main^exception^ that^ the^candidate^oligonucleotide^molecules^generated^ in^step^b)^ are^not^screened^for^their^ability^to^induce^a^reduction^of^the^titer^of^the^respiratory^virus^of^ interest^in^lung^tissue^but^rather^in^“conventional”^cell^culture.^Accordingly,^the^candidate^oli- gonucleotide^ molecules^ and^ the^ respiratory^ virus^ of^ interest^ are^ introduced^ as^ described^ herein^into^a^cell^in^culture^rather^than^into^a^lung^cut^slice.^The^cell^in^cell^culture^may^be^any^ suitable^cell^line^that^ is^susceptible^to^ infection^by^the^respiratory^virus^of^ interest.^In^some^ embodiments,^the^cell^may,^e.g.,^be^a^LLCMK2^cell^cultivated^in^a^suitable^cell^culture^medium^ such^as,^e.g.,^DMEM^supplemented^with^10%^FCS^+^Penicillin / Streptomycin.^Other^cells^that^ may^be^used^in^the^^“second^screening^step”^^include,^e.g.,^Calu3^cells^cultivated^in^a^suitable^ cell^culture^medium^such^as,^e.g.,^MEM^supplemented^with^10%^FCS^+^Penicillin / Streptomy- cin.^^ 17^ ^ As^described^herein,^the^candidate^oligonucleotide^molecule^can^be^introduced^into^the^cells^ by^means^of^transfection,^preferably^by^lipofection,^in^particular^when^the^candidate^oligonu- cleotide^molecule^is^an^siRNA^or^ASO,^whereas,^when^the^candidate^oligonucleotide^mole- cule^is^an^shRNA / shRNAmiR,^it^is^preferably^introduced^into^the^cell^by^means^of^a^suitable^ expression^vector^such^as^a^viral^vector.^ After^introduction^of^the^candidate^oligonucleotide^molecules,^the^cell^is^preferably^incubated^ at^a^temperature^of^32^to^40°C,^e.g.,^of^32,^33,^34,^35,^36,^37,^38,^39^or^40°C,^preferably^at^a^ temperature^of^35°C.^ The^cell^is^next^infected^with^a^suitable^amount^of^the^respiratory^virus^of^interest,^according^ to^ standard^ infection^protocols.^For^ instance,^ the^cell^may^be^ infected^with^a^ virus^ titre^ of^^ 2*102-^2*106,^e.g.,^of^2*102-^2*105,^2*103-^2*106,^^2*102-^2*104,^2*103-^2*105,^2*104-^2*106^or^of^2*102,^2*103,^2*104,^2*105^or^2*106^focus^forming^units^(FFU)^in^200µL.^Infection^of^the^cell^ with^the^respiratory^virus^of^interest^may^take^place^about^12-96^hours^,^e.g.^about^12,^24,^36,^ 48,^60,^72,^84^or^96^hours^after^introduction^of^the^candidate^oligonucleotide^molecules.^Pref- erably,^the^cell^is^infected^with^the^respiratory^virus^of^interest^24-84^hours^after^introduction^ of^the^candidate^oligonucleotide^molecules.^The^time^between^the^introduction^of^the^candi- date^oligonucleotide^molecules^and^viral^infection^may^depend^on^the^means^for^introducing^ the^candidate^oligonucleotide^molecule^ into^the^cell.^ If^the^candidate^oligonucleotide^mole- cules^are^ transduced^with^a^viral^vector^into^ the^cell,^ infection^with^ the^ respiratory^virus^of^ interest^may^preferably^be^done^about^72^hours^after^introduction^of^the^candidate^oligonu- cleotide^molecules^ to^ provide^ sufficient^ time^ for^ the^ expression^ and^ endonucleolytic^ pro- cessing^of^the^candidate^oligonucleotide^molecules.^If,^in^contrast,^the^candidate^oligonucle- otide^molecules^are^transfected,^e.g.,^as^siRNAs^or^ASOs^into^the^cell,^infection^with^the^res- piratory^virus^of^interest^may^already^be^done^about^24^hours^after^introduction^of^the^candi- date^oligonucleotide^molecule.^The^skilled^person^will^have^no^problem^to^determine^the^ideal^ time^point^ for^ infecting^ the^cell^with^ the^ respiratory^virus^of^ interest^within^ the^ limits^of^ the^ present^specification.^In^some^embodiments,^it^may^be^even^advantageous^to^introduce^the^ candidate^oligonucleotide^into^the^cell^and^simultaneously^infect^the^cell^with^the^respiratory^ virus.^Alternatively,^the^cell^may^first^be^infected^with^the^respiratory^virus^of^interest^before^ introducing^the^candidate^oligonucleotide^molecule.^For^instance,^the^candidate^oligonucleo- tide^molecules^may^be^introduced^into^the^cell^about^12-36^hours,^preferably^24^hours,^after^ it^has^been^infected^with^the^respiratory^virus^of^interest.^ After^introducing^the^candidate^oligonucleotide^molecule^and^infecting^the^cell^with^the^respir- atory^virus^of^interest,^culture^supernatant^is^obtained^from^the^cultured^cell^at^a^first^time^point^ and^at^ least^ a^ second^ time^point^ and^ the^virus^ titre^ in^each^supernatant^ is^determined^as^ previously^described.^ 18^ ^ At^the^end^of^the^“second^screening^step”,^those^candidate^oligonucleotide^molecules^causing^ a^reduction^of^the^viral^titre^in^the^supernatant^of^the^second^time^point^by^more^than^50%,^ preferably^by^more^than^60%,^70%^or^80%^compared^to^the^viral^titre^in^the^supernatant^of^ the^first^time^point^may^be^considered^suitable^for^use^in^an^oligonucleotide-based^therapeutic^ for^treating^an^infection^with^the^respiratory^virus^of^interest.^ The^screening^method^of^the^present^invention^may^comprise^either^only^the^“first^screening^ step”^or^only^the^“second^screening^step”^as^disclosed^herein^after^step^b).^Preferably,^how- ever,^the^method^comprises^both^the^“first”^and^the^“second^screening^steps”^as^described^ herein,^wherein^these^two^screening^steps^can^be^performed^in^any^order^after^step^b)^of^the^ method^of^the^invention.^Preferably,^in^such^an^embodiment,^a^candidate^oligonucleotide^mol- ecule^may^be^considered^suitable^for^use^in^an^oligonucleotide-based^therapeutic^for^treating^ an^ infection^with^the^ respiratory^virus^of^ interest^only^if^ it^achieves^the^desired^knockdown^ efficiency^in^the^“first^screening^step”^and^the^desired^reduction^in^viral^titre^ in^the^“second^ screening^step”,^as^defined^herein.^However,^in^some^embodiments,^a^candidate^oligonucle- otide^molecule^may^also^be^considered^for^use^in^an^oligonucleotide-based^therapeutic^for^ treating^an^infection^with^the^respiratory^virus^of^interest^even^if^it^only^“passes”^one^of^the^two^ screening^steps,^ i.e.,^even^ if^ it^only^achieves^the^desired^knockdown^efficiency^in^the^“first^ screening^step”^but^not^the^desired^reduction^in^viral^titre^in^the^“second^screening^step”^or^ vice^versa.^ It^can^be^advantageous^to^deliver^therapeutic^drugs^against^respiratory^diseases^in^the^form^ of^ aerosols,^ i.e.,^as^ fine^solid^particles^or^liquid^droplets^suspended^or^dispersed^ in^air^ that^ can^be^inhaled^by^a^patient^for^direct^ transport^to^the^lungs.^Accordingly,^ in^some^embodi- ments^of^the^screening^method^of^the^invention,^it^is^preferred^to^also^screen^the^candidate^ oligonucleotide^molecules^generated^in^step^b)^of^the^method^of^the^invention^for^their^knock- down^efficiencies^and / or^their^ability^to^induce^a^reduction^of^the^titer^of^^the^respiratory^virus^ of^interest^after^the^candidate^oligonucleotide^molecules^have^been^aerosolized.^Aerosoliza- tion^refers^to^the^process^of^converting^a^physical^substance^into^the^form^of^solid^particles^or^ liquid^droplets^small^and^light^enough^to^be^carried^on^the^air^i.e.^into^an^aerosol.^^ Thus,^in^some^embodiments,^the^screening^method^of^the^invention^may^comprise^an^addi- tional^step^after^step^b)^comprising^aerosolzing^the^candidate^oligonucleotide^molecules^gen- erated^in^step^b),^wherein^the^aerosolized^candidate^oligonucleotide^molecule^is^subsequently^ subjected^to^one^or^both^of^the^herein^disclosed^screening^steps^(i.e.,^the^“first^screening^step”^ and / or^the^“second^screening^step”).^ The^skilled^person^will^be^aware^of^various^methods^that^can^be^used^to^aerosolize^an^oligo- nucleotide^molecule^such^as^an^ASO^or^an^siRNA.^For^ instance,^aerosols^containing^liquid^ droplets^comprising^the^candidate^oligonucleotide^molecules^can^be^obtained^by^a^process^ 19^ ^ known^as^nebulization.^For^nebulization,^the^candidate^oligonucleotide^molecules^are^prefer- ably^first^introduced^into^a^suitable^delivery^vehicle^such^as,^e.g.,^an^LNP^or^are^associated^ with^cationic^polymers^or^ lipophilic^anchors,^as^described^herein,^before^being^suspended^ into^a^liquid^such^as,^e.g.,^water.^Nebulization^of^this^suspension^into^aerosols^containing^the^ candidate^oligonucleotide^molecules^can^then^be^achieved^using^any^nebulizer^device^known^ in^the^art,^e.g.,^a^jet^nebulizer,^a^vibrating^mesh^nebulizer^or^an^ultrasonic^nebulizer.^ Alternatively,^aerosolization^of^the^candidate^oligonucleotide^molecule^can^be^achieved^using,^ e.g.,^a^dry^powder^inhaler.^In^this^case,^the^resulting^aerosols^do^not^comprise^liquid^droplets^ but^a^fine^powder^formed^by^the^candidate^oligonucleotides.^Also^in^such^an^embodiment,^the^ candidate^oligonucleotide^molecule^is^preferably^first^introduced^into^a^suitable^delivery^vehi- cle^such^as,^e.g.,^ an^LNP^or^ is^associated^with^cationic^polymers^or^ lipophilic^anchors,^as^ described^herein,^prior^to^aerosolization.^However,^in^contrast^to^nebulization,^the^candidate^ oligonucleotide^molecule^is^not^subsequently^suspended^in^a^liquid,^but^filled^directly^into^the^ dry^powder^inhaler^for^aerosolization.^ After^ aerosolization,^ the^average^ droplet^ or^ particle^ diameter^within^ the^obtained^aerosols^ comprising^the^candidate^oligonucleotide^molecule^preferably^is^less^than^10^µm.^Particles^of^ more^than^10^μm^in^diameter^are^less^desired^because^they^are^more^likely^to^deposit^in^the^ mouth^and^throat^and^thus^might^not^reach^the^lungs^of^a^patient.^Preferably,^ the^average^ droplet^or^particle^diameter^is^even^less^than^5^µm,^because^particles^smaller^than^5^μm^in^ diameter^were^found^to^deposit^more^frequently^in^the^lower^airways^and^are^thus^particularly^ suitable^for^pharmaceutical^aerosols.^The^average^droplet^or^particle^diameter^may^also^be^ less^than^4^µm,^less^than^3^µm,^less^than^2^µm^or^even^less^than^1^µm.^ Preferably,^the^aerosolized^candidate^oligonucleotide^molecules^are^subsequently^collected^ by^ any^ suitable^ method^ or^ device^ known^ to^ the^ skilled^ person^ (described^ e.g.,^ in^ https: / / edoc.ub.uni-muenchen.de / 17279 / 1 / Hertel_Sebastian.pdf),^ and^ introduced^ into^ the^ cells^used^in^any^of^the^screening^steps^disclosed^herein.^ The^present^invention^further^provides^a^method^of^preparing^an^oligonucleotide-based^ther- apeutic^for^treating^an^infection^of^a^respiratory^virus^of^interest.^^ The^method^comprises^steps^of^selecting^a^candidate^oligonucleotide^molecule^capable^of^ inducing-mediated^immunity^against^a^respiratory^virus^using^the^herein^described^screening^ method^and^providing^a^vector^comprising^or^encoding^said^candidate^oligonucleotide^mole- cule^to^obtain^the^oligonucleotide^-based^therapeutic^for^treating^a^respiratory^virus^of^interest.^ As^already^described^herein,^the^candidate^oligonucleotide^molecule^selected^by^the^screen- ing^ method^ according^ to^ the^ invention^ can^ be^ provided^ as^ an^ siRNA,^ an^ shRNA,^ an^ shRNAmiR^(also^referred^herein^as^artificial^miRNA)^or^an^ASO.^For^instance,^the^candidate^ 20^ ^ oligonucleotide^molecule^may^be^selected^based^on^its^efficiency^to^reduce^viral^load^in^cells^ during^the^second^screening^step^of^the^screening^method^according^to^the^invention.^ In^the^context^of^the^invention,^the^term^“vector”^refers^to^any^means^suitable^for^delivering^ an^RNA^molecule^capable^of^inducing^RNAi^or^an^ASO^to^a^target^cell^or^tissue,^in^particular^ a^cell^of^the^respiratory^tract,^herein^also^referred^to^as^airway^cell.^The^choice^of^a^suitable^ vector^of^the^oligonucleotide-based^therapeutic^essentially^depends^on^whether^the^candidate^ oligonucleotide^molecule^is^to^be^provided^in^the^form^of^an^siRNA^or^ASO^or^in^the^form^of^ an^shRNA / shRNAmiR.^In^addition,^the^choice^of^vector^depends^on^whether^ the^candidate^ oligonucleotide^molecule^is^to^be^expressed^at^its^intended^site^of^action^or^is^provided^as^an^ in^vitro-produced^oligonucleotide^molecule^that^needs^to^be^transported^to^its^target^site.^For^ instance,^ if^ the^ candidate^ oligonucleotide^ molecule^ is^ provided^ as^ an^ shRNA^ or^ an^ shRNAmiR,^the^vector^preferably^is^an^expression^vector^encoding^the^candidate^oligonucle- otide^molecule,^as^described^herein,^i.e.^it^may^be^a^minicircle,^a^plasmid,^a^cosmid,^a^phage,^ a^virus^or^an^artificial^chromosome.^Preferably,^the^expression^vector^is^a^viral^vector,^e.g.,^ an^adenoviral^vector,^an^adeno-associated^vector^(AAV),^or,^most^preferably^a^retroviral,^e.g.,^ lentiviral^vector.^^ The^expression^vector^comprises^an^expression^cassette,^i.e.^the^necessary^elements^ that^ permit^transcription^of^a^cargo^nucleic^acid^sequence^into^the^candidate^oligonucleotide^mol- ecule,^e.g.^the^shRNA^or^shRNAmiR,^such^as^a^suitable^promoter.^In^one^embodiment,^the^ expression^vector^may^comprise^a^cargo^sequence^that^encodes^only^a^single^candidate^oli- gonucleotide^molecule,^e.g.^a^single^shRNA^or^shRNAmiR,^selected^by^the^screening^method^ of^ the^ invention.^Alternatively,^ the^expression^vector^may^comprise^a^cargo^sequence^that^ encodes^a^plurality^of^different^candidate^oligonucleotide^molecules,^e.g.,^a^plurality^of^differ- ent^shRNAs^or^shRNAmiRs,^selected^by^the^screening^method^of^the^invention.^This^way,^it^ is^ possible^ to^express^multiple^different^candidate^oligonucleotide^molecules^with^ different^ target^sequences^from^a^single^expression^vector.^ A^person^skilled^ in^the^art^will^be^able^to^design^suitable^viral^vectors^to^comprise^a^cargo^ DNA^sequence^encoding^one^or^more^candidate^oligonucleotide^molecules,^e.g.^one^or^more^ shRNAs^or^shRNAmiRs^based^on^general^knowledge^and^the^prior^art.^For^example,^a^stretch^ of^cargo^DNA,^i.e.,^a^transgene,^encoding^the^one^or^more^candidate^oligonucleotide^mole- cules^may^be^operably^ linked^ to^ a^ suitable^ promoter^ and^ inserted^ into^ the^ viral^ vector^of^ choice.^The^promoter^used^to^express^the^one^or^more^oligonucleotide^molecules^may^be^any^ constitutively^active^promoter^known^in^the^art,^e.g.,^a^Rous^sarcoma^virus^(RSV)^promoter,^ a^human^cytomegalovirus^(CMV)^promoter,^an^HIV^promoter^or^a^eukaryotic^promoter^such^ as^e.g.,^EF1a,^PGK1,^UBC,^or^human^beta^actin.^Alternatively,^the^promoter^may^be^an^in- ducible^promoter^ that^only^drives^expression^of^ the^one^or^more^candidate^oligonucleotide^ molecules^in^the^presence^or^absence^of^a^certain^stimulus.^For^example,^the^inducible^pro- moter^may^be^a^Tet-regulated^promoter,^i.e.,^it^may^comprise^a^tetracycline^response^element^ 21^ ^ (TRE)^ that^ can^ be^bound^by^a^ tetracycline^ transactivator^ (tTA)^protein^ in^ the^presence^of^ tetracycline^or^an^analogue^thereof,^e.g.,^doxycycline.^The^promoter^may^also^be^a^cell-spe- cific^promoter^which^only^drives^expression^in^a^particular^type^of^cell,^e.g.,^an^airwaycell.^^ In^a^preferred^embodiment,^the^vector^furthermore^is^a^cell-specific^vector^capable^of^targeting^ the^ one^ or^ more^ candidate^ oligonucleotide^ molecules,^ e.g.^ the^ one^ or^ more^ shRNAs^ or^ shRNAmiRs,^to^a^cell^or^a^tissue^of^interest,^e.g.,^to^cells^of^the^respiratory^tract.^Accordingly,^ if^the^vector^is^a^viral^vector^such^as^a^lentiviral^vector,^the^assembled^viral^vector^is^preferably^ pseudotyped,^e.g.,^with^one^or^more^different^viral^envelope^glycoproteins^or^a^capsid^to^fa- cilitate^binding^of^the^vector^to^the^surface^of^the^cell^of^interest,^e.g.,^an^airway^cell.^The^term^ pseudotyping^refers^to^the^generation^of^viral^vectors^that^carry^foreign^viral^envelope^glyco- proteins^on^their^surface^or^are^encapsulated^in^a^capsid^of^a^different^virus^or^viral^serotype^ to^specifically^target^only^particular^cell^types^of^interest.^The^viral^vector^particles^are^usually^ assembled^inside^suitable^packaging^cells^known^in^the^art^according^to^standard^laboratory^ techniques^before^they^are^administered^to^the^subject.^^ The^expression^vector^used^for^delivering^ the^candidate^oligonucleotide^molecule,^e.g.^ the^ shRNA^or^shRNAmiR,^may^also^be^a^non-pathogenic,^genetically^modified^bacterium^or^yeast^ comprising^a^prokaryotic^or^eukaryotic^vector.^The^vector^comprises^a^DNA^molecule^encod- ing^ the^candidate^oligonucleotide^molecule^operably^ linked^ to^a^promoter^ that^controls^ex- pression^of^ said^candidate^oligonucleotide^molecule,^as^described^above.^The^non-patho- genic^bacterium^is^preferably^invasive,^i.e.,^it^is^capable^of^entering^a^host^cell^and^may^be^ derived,^e.g.,^from^a^non-pathogenic^strain^of^Escherichia^coli,^Listeria,^Yersinia,^Rickettsia,^ Shigella,^ Salmonella,^ Legionella,^Chlamydia,^ Brucella,^Neisseria,^ Burkolderia,^ Bordetella,^ Borrelia,^Coxiella,^Mycobacterium,^Helicobacter,^Staphylococcus,^Streptococcus,^Vibrio,^Lac- tobacillus,^Porphyromonas,^Treponema,^or^Bifidobacteriae.^Preferably,^the^prokaryotic^or^eu- karyotic^vector^is^specifically^designed^for^targeted^delivery^to^a^cell^of^ interest,^e.g.,^by^ex- pressing^a^ligand^on^its^surface^that^can^be^recognized^by^a^receptor^on^the^target^cell,^e.g.^ an^airway^cell.^ However,^ in^ some^ embodiments,^ it^ may^ also^ be^ advantageous^ not^ to^ express^ shRNAs / shRNAmiRs^in^situ^from^an^expression^vector.^Rather,^the^candidate^oligonucleotide^ molecule^ in^ the^ form^of^an^shRNA / shRNAmiR^may^ first^be^produced^ in^vitro^before^being^ packaged^into^a^suitable^delivery^vehicle^for^direct^delivery^to^the^target^cell.^The^use^of^such^ transport^vehicles^as^vectors^is^also^preferred^in^cases^where^the^candidate^oligonucleotide^ molecule^is^not^provided^as^shRNA^or^shRNAmiR,^but^as^siRNA^or^ASO,^since^siRNAs^or^ ASOs^are^typically^not^expressed^in^situ^from^an^expression^vector^at^the^cell^of^interest,^but^ are^provided^as^constructs^generated^in^vitro.^^ A^person^skilled^ in^the^art^ is^aware^of^various^types^of^vectors^that^may^serve^as^delivery^ vehicles^for^in^vitro^generated^oligonucleotide^molecules^such^as^ASOs,^siRNAs^or^in^vitro- 22^ ^ transcribed^shRNAs / shRNA / miRs^and^which^are^capable^of^selectively^targeting^the^oligonu- cleotide^to^a^cell^of^interest.^In^a^preferred^embodiment,^the^vector^may^e.g.,^comprise^a^lipid^ membrane^that^encapsulates^the^candidate^oligonucleotide^molecule.^For^example,^the^vec- tor^may^be^a^liposomal^or^an^exosomal^carrier^comprising^the^candidate^oligonucleotide^mol- ecule^as^a^cargo^and,^optionally,^expressing^a^targeting^moiety^on^its^surface.^In^a^particularly^ preferred^embodiment,^the^vector^is^a^lipid^nanoparticle^(LNP)^which^typically^has^a^size^of^ about^10^to^1000^nanometers.^The^LNP^may^optionally^comprise^various^modifications,^in- cluding^variations^of^cholesterol,^lipid-anchored^polyethanilglycol,^structural^lipids^and^ioniza- ble^lipids.^^The^vector^may^however^also^be^a^bacterial^minicell.^These^types^of^carriers^have^ the^ability^to^entrap^both^hydrophilic^therapeutic^agents^within^their^central^aqueous^core^or^ lipophilic^drugs^within^their^bilayer^compartment.^The^targeting^moiety^may^be,^e.g.,^a^cell- specific^antibody^or^ligand^such^as,^e.g.,^a^small^peptide,^polysaccharide^or^nucleic^acid^that^ can^be^bound^by^a^receptor^expressed^on^ the^surface^of^the^cell^of^ interest,^e.g.,^a^airway^ cell.^Use^of^such^a^liposomal,^exosomal^or^minicell^vector^has^the^benefit^of^allowing^for^fusion^ of^the^vector^with^the^lipid^membrane^of^the^cell^of^interest^e.g.,^during^endocytosis^to^allow^ for^the^release^of^candidate^oligonucleotide^molecule^into^the^cytoplasm^of^the^cell.^^ In^another^embodiment,^the^vector^may^be^a^polymeric^micro-^or^nanoparticle.^The^micro-^or^ nanoparticle^may^be^formed^of^any^physiologically^acceptable^polymeric^material^known^in^ the^art,^e.g.,^it^may^comprise^a^biopolymer^such^as^polysaccharide^selected^from^the^group^ comprising^cellulose,^alginate^or^chitosan.^The^vector^may^also^be^a^nanocarrier^comprising,^ e.g.,^a^hyperbranched^dendritic^polyglycerol^polymer.^Such^a^nanocarrier^ is^an^amphiphilic^ unimolecular^nanocarrier^of^dendritic^structure,^and^it^comprises^a^dendritic^core^and^at^least^ one^shell.^Dendrimers^may^also^be^used.^Further^alternative^ vectors^ known^ to^ the^skilled^ person^include^e.g.,^polyethylenimine^(PEI)-based^vectors.^ In^one^embodiment,^the^vector^releases^the^candidate^oligonucleotide^molecule^at^once,^after^ the^vector^has^reached^its^targeted^destination^inside^the^subject.^In^another^possible^embod- iment,^the^vector^may^be^configured^in^a^fashion^to^allow^for^a^sustained^release^of^the^can- didate^oligonucleotide^molecule^in^a^desired^period^of^time.^A^person^skilled^in^the^art^will^be^ able^to^choose^a^suitable^vector^from^the^art^depending^on^the^type^of^oligonucleotide^used,^ the^route^of^administration,^and^the^respective^target^cell^to^which^the^candidate^oligonucleo- tide^molecule^is^to^be^delivered.^In^addition,^based^on^the^known^state^of^the^art,^the^skilled^ person^will^be^able^to^further^modify^the^vector^for^optimized^targeted^delivery^and^release^of^ the^inhibitor.^ In^some^embodiments,^the^in^vitro-generated^candidate^oligonucleotide^molecule^may^also^ not^be^incorporated^into^a^delivery^vehicle^as^described^herein,^but^may^alternatively^be^la- belled^with^a^suitable^ lipid^anchor^ to^ improve^the^delivery^of^ the^oligonucleotide^candidate^ molecule,^ e.g.,^ by^ directly^ conjugating^ the^ in^ vitro^ produced^ ASO,^ siRNA^ or^ shRNA / shRNAmiR^to^lipid^residues^such^as^palmityl-^(also^referred^to^as^2′-O-hexadecyl^or^ 23^ ^ C16)^or^cholesterol-residues^as^described,^e.g.,^in^Raouane^et^al.,^2012^or^Brown^et^al.,^2022.^ Accordingly,^said^lipid^anchor^may^be^considered^herein^to^serve^as^a^vector^for^delivery^of^ the^oligonucleotide^candidate^molecule.^ The^oligonucleotide-based^therapeutic^produced^by^the^herein^described^method^is^prefera- bly^ formulated^ in^a^pharmaceutical^composition.^Accordingly,^ oligonucleotide-based^ thera- peutic^can^be^provided^in^a^formulation^with^one^or^more^excipients,^e.g.^with^a^pharmaceu- tically^acceptable^carrier.^ Using^the^herein^described^screening^method,^the^inventors^were^able^to^identify^oligonucle- otide^molecules^that^are^capable^of^inducing^RNAi-mediated^immunity^against^human^parain- fluenza^virus^or^of^ inducing^antisense-mediated^degradation^of^RNAs^derived^ from^human^ parainfluenza^virus.^Human^parainfluenza^virus^(HPIV)^is^a^respiratory^virus^that^can^cause^ severe^complications,^including^death,^especially^in^immunosuppressed^patients^(for^exam- ple,^stem^cell^ transplant^ recipients^or^ those^with^chronic^ lung^disease).^ It^ is^also^the^most^ common^cause^of^pseudocroup^in^children.^To^date,^there^is^neither^a^specific^therapy^nor^an^ effective^vaccine^against^HPIV.^^ HPIV^is^a^single-stranded^RNA^virus^of^the^Paramyxoviridae^family^with^a^negative^sense^(-)^ RNA^genome^encompassing^about^15,000^nucleotides.^HPIV^has^six^essential^genes^desig- nated^as^L,^HN,^M,^PC,^F,^NP,^which^all^constitute^potential^targets^for^RNAi^mediating^agents.^ As^described^in^the^example^below,^the^inventors^followed^the^steps^of^the^herein^described^ screening^method^to^identify^numerous^novel^ target^sequences^within^both^the^(-)RNA^ge- nome^of^HPIV^and^the^(+)RNA^intermediate^formed^for^gene^expression^and^subsequently^ generated^a^series^of^candidate^oligonucleotide^molecules^expected^to^recognize^said^target^ sequences^and^thus^to^trigger^RNAi^or,^in^case^of^ASOs,^to^induce^antisense-mediated^deg- radation^or^blockage^of^the^viral^RNA.^^ Thus,^the^present^invention^further^provides^an^oligonucleotide^molecule^capable^of^targeting^ an^RNA^derived^from^HPIV,^wherein^the^oligonucleotide^molecule^is^^ a)^ an^ RNA^ molecule^ capable^ of^ inducing^ RNAi^ comprising^ a^ nucleic^ acid^ se- quence^having^at^least^90%,^e.g.,^at^least^95%^or^100%^sequence^identity^to^SEQ^ID^ NO:^1^to^56;^or^ b)^ an^ASO^comprising^a^nucleic^acid^sequence^that^corresponds^to^at^least^a^sub- sequence^of^10^or^more^nucleotides^of^any^of^SEQ^ID^NO:^1^to^56.^ In^the^context^of^the^invention,^the^term^“RNA^derived^from^HPIV”^refers^either^to^the^negative- sense^(-)^RNA^HPIV^genome^or^the^(+)^RNA^intermediate^produced^by^HPIV^for^gene^ex- pression.^ 24^ ^ The^term^“targeting”^is^herein^understood^to^mean^that^the^oligonucleotide^molecules^of^the^ invention^are^able^to^bind,^i.e.,^to^“hybridize”^to^a^target^RNA^derived^from^HPIV^via^comple- mentary^Watson–Crick^base^pairing^as^defined^herein.^ SEQ^ID^NO:^1^to^56^relate^to^ribonucleic^acid^sequences.^However,^it^is^to^be^understood^that^ a^nucleic^acid^sequence^that^corresponds^to^at^least^SEQ^ID^NO:^1^to^56^or^subsequences^ thereof^may^be^a^deoxyribonucleic^acid^sequence^or^ribonucleic^acid^sequence.^It^may^even^ comprise^both^deoxyribonucleotides^and^ribonucleotides^and / or^modified^nucleotides,^e.g.,^in^ case^the^ASO^is^a^chimeric^gapmer,^as^described^below.^^^^ In^some^embodiments,^the^oligonucleotide^molecule^of^ the^ invention^is^a^synthetic^siRNA.^ The^synthetic^siRNA^is^typically^provided^as^an^RNA^duplex^of^20^to^27^base^pairs^(bp)^con- sisting^of^a^sense^and^a^complementary^antisense^strand.^The^antisense^strand^of^the^siRNA^ duplex^comprises^a^nucleic^acid^sequence^having^at^least^90%,^preferably^at^least^95%,^more^ preferably^100%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^56.^Alternatively,^the^antisense^ strand^of^the^siRNA^duplex^may^also^consist^of^a^nucleic^acid^sequence^having^at^least^90%,^ preferably^at^least^95%,^more^preferably^100%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^ 56.^ In^other^words,^ the^siRNA^antisense^strand^may^comprise^or^consist^of^a^nucleic^acid^ sequence^that^comprises^up^to^2,^i.e.,^0,^1^or^2^nucleotide^substitutions^compared^to^any^of^ SEQ^ID^NO:^1^to^56.^Once^the^siRNA^has^been^introduced^into^a^cell,^the^antisense^strand^of^ the^siRNA^duplex^serves^as^the^guide^strand,^which^is^incorporated^into^RISC^to^target^the^ complex^ to^ the^HPIV-derived^RNA^for^RNAi.^In^different^embodiments,^ the^oligonucleotide^ molecule^may^also^be^only^one^strand^of^an^siRNA^duplex,^i.e.^it^may^only^be^the^antisense^ strand^comprising^or^consisting^of^a^nucleic^acid^sequence^having^at^least^90%,^preferably^at^ least^95%,^more^preferably^100%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^56.^In^such^a^ scenario,^the^RNA^molecule^capable^of^inducing^RNAi-mediated^immunity^against^HPIV^may^ also^consist^of^a^nucleic^acid^sequence^having^at^least^90%,^preferably^at^least^95%,^more^ preferably^100%^sequence^identity^any^of^SEQ^ID^NO:^1^to^56.^ In^alternative^embodiments,^the^RNA^molecule^capable^of^inducing^RNAi^may^be^an^shRNA,^ whereas^a^nucleic^acid^sequence^having^at^least^90%,^preferably^at^least^95%,^more^prefer- ably^100%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^56^forms^a^part^of^the^stem^of^said^ shRNA.^Accordingly,^when^the^RNA^molecule^ is^an^shRNA,^it^necessarily^comprises^addi- tional^nucleotides^besides^the^nucleic^acid^sequence^having^at^least^90%,^preferably^at^least^ 95%,^more^preferably^100%^sequence^identity^toany^of^SEQ^ID^NO:^1^to^56^to^facilitate^proper^ folding^of^the^RNA^molecule^into^the^stem^loop^structure^of^an^shRNA.^Preferably,^the^shRNA^ is^embedded^into^a^miRNA^backbone,^also^referred^to^as^miRNA^framework,^as^described^ herein,^ e.g.,^ the^ framework^of^miR30a,^miR223,^miR144^or^miR33.^ Accordingly,^ the^RNA^ molecule^may^also^be^an^shRNAmiR.^ 25^ ^ The^skilled^person^will^be^aware^that^neither^siRNAs^nor^miRNAs^or^shRNA / shRNamiR^must^ be^fully^complementary^ to^ their^ target^sequences^to^effectively^suppress^protein^synthesis,^ i.e.,^he^or^she^will^know^that^some^mismatches^are^admissible.^Thus,^if^the^herein^disclosed^ siRNA^antisense^strand^or^the^guide^sequence^of^the^shRNA / shRNamiR^comprises^one^or^ two^substitutions^compared^to^any^of^SEQ^ID^NO:^1^to^56,^it^binds^to^its^target^sequence^with^ imperfect^complementarity^but^can^nevertheless^be^a^potent^RNAi^trigger^that^specifically^and^ effectively^induces^cleavage^of^the^target^RNA^or^inhibits^its^translation^into^protein.^However,^ in^preferred^embodiments,^the^nucleotides^corresponding^to^positions^2-8^of^SEQ^ID^NO:^1^ to^56^are^not^substituted^in^the^herein^disclosed^siRNAs^or^shRNAs / shRNAmiRs,^as^these^ positions^form^the^so-called^“seed”^region^critically^for^the^specificity^of^an^RNAi^trigger.^More- over,^multiple^studies^^(e.g.,^Du^et^al.,^2005;^Ahmed^and^Raghava,^2011)^suggest^that^mis- matches^affecting^positions^4^-12^in^siRNAs^significantly^reduce^RNAi^efficacy,^whereas^mis- matches^at^positions^1,^2,^3,^18^and^19,^in^particular^at^positions^1^and^19,^appear^to^affect^ RNAi^efficacy^only^weakly^or^not^at^all^ (Ahmed^and^Raghava,^2011).^ In^view^of^this,^ if^ the^ RNA^molecule^of^the^invention^capable^of^inducing^RNAi^comprises^or^consists^of^a^nucleic^ acid^sequence^having^90%^or^95%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^56,^it^prefer- ably^does^not^comprise^a^substitution^at^a^position^that^correspond^to^positions^2-12^of^SEQ^ ID^NO:^1^to^56.^It^may,^e.g.,^comprise^a^substitution^at^a^position^that^corresponds^to^position^ 1^and / or^position^19^of^SEQ^ID^NO:^1^to^56.^^An^RNA^molecule^of^the^invention^capable^of^ inducing^RNAi^may^also^comprise^or^consist^of^a^nucleic^acid^sequence^that^is^shorter^than^ the^sequences^of^SEQ^ID^NO:^1^to^56,^e.g.,^it^may^comprise^or^consist^only^of^a^subsequence^ spanning^20^or^21^nucleotides,^e.g.,^the^first^20^or^21^nt,^of^any^of^SEQ^ID^NO:^1^to^56.^ The^RNA^molecule^capable^of^ inducing^RNAi^of^ the^ invention^may^comprise^one^or^more^ chemical^modifications^that^have^a^stabilizing^effect,^in^particular,^in^case^the^RNA^molecule^ is^an^siRNA.^For^instance,^the^RNA^molecule^may^comprise^a^modification^in^its^ribose^sugar^ backbone,^e.g.,^a^2’-O^modification^such^as^2′-OMe,^2′-F,^2′-O-methoxyethyl^(2′-MOE)^or^2′- O-guanidinopropyl^(2′-O-GP).^ Additional^modification^that^may^be^incorporated^in^the^RNA^molecule^of^the^invention^may^ include,^e.g.,^5-fluoro-2′-deoxyuridine^and^2′-O-methyl^phospshorodithioate^moieties.^ The^aforementioned^modifications^as^well^as^others^that^may^be^used^in^the^herein^disclosed^ RNA^molecules^are^well^known^to^the^skilled^person^and^are^summarized^e.g.,^in^Selvam^et^ al.,^2017.^Modifications^may^also^be^added^to^improve^siRNA^delivery^as^described^above.^ For^ instance,^^one^or^more^nucleotides^in^the^passenger^RNA^strand^of^ the^siRNA^duplex^ may^be^conjugated^to^a^suitable^lipophilic^moiety^or^anchor^such^as,^e.g.,^palmityl-^(also^re- ferred^ to^ as^2′-O-hexadecyl^ or^C16)^ or^ cholesterol-residues^ as^described^above.^Palmityl^ modifications^of^siRNAs^are^known^to^the^skilled^person^due^to,^e.g.^Brown^et^al.,^2022.^ 26^ ^ Using^the^method^of^the^invention,^ the^present^inventors^could^show^that^at^least^twelve^of^ the^56^RNA^molecules^disclosed^herein^were^able^to^target^an^RNA^derived^from^HPIV^and^ induce^a^reduction^of^the^HPIV^titer^in^human^PCLS^when^they^were^provided^either^as^siR- NAs^or^shRNA / shRNAmiR.^The^RNA^molecules^that^most^effectively^induced^RNAi-mediated^ immunity^against^HPIV^comprised^a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO.^1,^2,^3,^9,^ 12,^14,^15,^17,^18,^22,^23^and^29.^Accordingly,^in^a^preferred^embodiment,^the^RNA^molecule^ of^the^invention^capable^of^inducing^RNAi^comprises^a^nucleic^acid^sequence^of^any^of^SEQ^ ID^NO.^1,^2,^3,^9,^12,^14,^15,^17,^18,^22,^23^and^29.^As^described^herein,^the^RNA^molecule^ may^also^comprises^a^nucleic^acid^sequence^comprising^one^or^two^substitutions^compared^ to^any^of^SEQ^ID^NO:^1,^2,^3,^9,^12,^14,^15,^17,^18,^22,^23^and^29.^Accordingly,^ the^RNA^ molecule^capable^of^ inducing^RNAi^may^also^comprise^a^nucleic^acid^sequence^having^at^ least^90%,^or^at^least^95%^sequence^identity^to^any^of^SEQ^ID^NO:^1,^2,^3,^9,^12,^14,^15,^17,^ 18,^22,^23^and^29.^If^the^RNA^molecule^capable^of^inducing^RNAi^is^an^siRNA,^its^guide^strand^ may^also^consist^a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO.^1,^2,^3,^9,^12,^14,^15,^17,^18,^ 22,^23^and^29^or^a^nucleic^acid^sequence^having^at^ least^90%,^or^at^ least^95%^sequence^ identity^thereto.^Preferably,^if^the^RNA^molecule^of^the^invention^is^an^siRNA,^it^consists^of^a^ guide^strand^and,^optionally,^a^complementary^passenger^strand^according^to^table^7^of^the^ example^below.^It^thus^may^consist^of^a^guide^strand^having^a^nucleic^acid^sequence^of^any^ of^SEQ^ID^NO:^169,^171,^173,^175^or^177^and,^optionally,^a^corresponding^complementary^ passenger^strand^having^a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO:^170,^172,^174,^176^ or^178,^respectively.^The^siRNA^may^alternatively^also^consist^of^a^guide^strand^with^a^nucleic^ acid^sequence^having^at^least^90%,^or^at^least^95%^sequence^identity^to^any^of^SEQ^ID^NO:^ 169,^171,^173,^175^or^177^and,^optionally,^a^corresponding^complementary^passenger^strand^ with^a^nucleic^acid^sequence^having^at^least^90%,^or^at^least^95%^sequence^identity^to^any^ of^SEQ^ID^NO:^170,^172,^174,^176^or^178,^respectively.^In^other^words,^the^siRNA^antisense^ or^guide^strand^may^consist^of^a^nucleic^acid^sequence^that^comprises^up^to^2,^i.e.,^0,^1^or^2^ nucleotide^substitutions^compared^to^any^of^SEQ^ID^NO:^169,^171,^173,^175^or^177,^whereas,^ optionally,^the^corresponding^complementary^passenger^strand^may^consist^of^a^nucleic^acid^ sequence^that^comprises^up^to^2,^i.e.,^0,^1^or^2^nucleotide^substitutions^compared^to^any^of^ SEQ^ID^NO:^170,^172,^174,^176^or^178,^respectively.^However,^the^^guide^strand^of^the^siRNA^ preferably^does^not^comprise^a^substitution^at^a^position^that^corresponds^to^positions^2-12^ of^any^of^SEQ^ID^NO:^169,^171,^173,^175^or^177.^It^may,^e.g.,^comprise^a^substitution^at^a^ position^that^corresponds^to^position^1^and / or^position^19^of^SEQ^ID^NO:^169,^171,^173,^175^ or^177.^^The^guide^strand^of^the^siRNA^of^the^invention^may^also^consist^of^a^nucleic^acid^ sequence^that^is^shorter^than^the^sequences^of^SEQ^ID^NO:^169,^171,^173,^175^or^177,^e.g.,^ it^may^comprise^or^consist^only^of^a^subsequence^spanning^25^or^26^nucleotides,^e.g.,^the^ first^25^or^26^nt,^of^any^of^SEQ^ID^NO:^:^169,^171,^173,^175^or^177.^ In^a^particular^preferred^embodiment,^the^RNA^molecule^capable^of^inducing^RNAi^comprises^ a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO:^1,^18^and^23^or^a^nucleic^acid^sequence^having^ 27^ ^ at^least^90%,^or^at^least^95%^sequence^identity^thereto,^as^these^three^candidates^turned^out^ to^ be^ particularly^ effective^ for^ inducing^ RNAi-mediated^ immunity^ against^ HPIV^ in^ human^ PCLS.^SEQ^ID^NO:^1,^18^and^23^are^all^complementary^to^a^target^sequence^in^the^(+)RNA^ intermediate^produced^by^HPIV.^As^can^be^seen^in^the^example^below,^RNA^molecules^com- prising^either^SEQ^ID^NO:^1^or^23^effectively^reduced^HPIV^titres^in^PCLS^ independent^of^ whether^ they^were^provided^as^siRNAs^or^as^shRNAmiRs^with^a^miR-30a^ framework.^By^ comparison,^ the^RNA^molecule^comprising^a^nucleic^acid^of^SEQ^ID^NO:^18^ is^preferably^ provided^as^an^siRNA^to^effectively^mediate^RNAi-induced^ immunity^against^HPIV^ in^ lung^ tissue.^ Therefore,^if^the^RNA^molecule^is^an^siRNA,^it^preferably^consists^of^a^guide^strand^having^a^ nucleic^acid^sequence^of^any^of^SEQ^ID^NO:^169,^173^or^175^or^a^nucleic^acid^sequence^ having^at^least^90%,^or^at^least^95%^sequence^identity^thereto^and,^optionally,^a^correspond- ing^complementary^passenger^strand^having^a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO:^ 170,^174,^or^176,^respectively,^or^a^nucleic^acid^having^at^least^90%,^or^at^least^95%^sequence^ identity^thereto.^ In^another^embodiment,^the^oligonucleotide^molecule^of^the^invention^may^be^an^ASO^com- prising^a^nucleic^acid^sequence^that^corresponds^to^at^ least^a^subsequence^of^10^or^more^ nucleotides^of^any^of^SEQ^ID^NO:^1^to^56.^^Said^subsequence^may^correspond^to^any^con- tinuous^stretch^of^at^least^10^nt^length^present^in^any^of^SEQ^ID^NO:^1^to^56.^For^instance,^ the^ASO^may^comprise^a^nucleic^acid^sequence^that^corresponds^to^at^least^the^first^or^final^ 10^nucleotides^of^any^of^SEQ^ID^NO:^1^to^56,^preferably,^the^final^10^nucleotides.^The^ASO^ may^also^comprise^a^nucleic^acid^sequence^ that^corresponds^to^a^subsequence^of^any^of^ SEQ^ID^NO:^1^to^56^that^is^longer^than^10^nt,^e.g.,^it^may^comprise^a^nucleic^acid^sequence^ that^corresponds^to^a^continuous^subsequence^spanning^11,^12,^13,^14,^15,^16,^17,^18,^19,^ 20^or^21^ nt^of^any^of^SEQ^ID^NO:^1^ to^56.^Preferably,^ the^ASO^comprises^a^nucleic^acid^ sequence^that^corresponds^to^at^ least^a^subsequence^of^12^or^more^nucleotides^of^any^of^ SEQ^ID^NO:^1^ to^56.^ ^ In^some^embodiments,^ the^ASO^may^also^comprise^a^nucleic^acid^ sequence^that^corresponds^to^the^complete^sequence^of^any^of^SEQ^ID^NO:^1^to^56^or^con- sists^thereof.^Preferably,^the^ASO^comprises^a^nucleic^acid^sequence^that^corresponds^to^at^ least^a^subsequence^of^10^or^more^continuous^nucleotides^of^any^of^SEQ^ID^NO:^1,^2,^3,^9,^ 12,^14,^15,^17,^18,^22,^23^and^29,^more^preferably^of^SEQ^ID^NO:^1,^18^and^23.^Particular^ preferred^ASOs^of^the^invention^comprise^a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO:^204,^ 205^or^206.^ The^ASOs^of^the^invention^may^be^DNA^ASOs.^^ The^ASOs^of^the^invention^may^comprise^a^variety^of^chemical^modifications^to^improve^their^ pharmacokinetic^properties^such^as^stability,^specificity^and^membrane^permeability,^and^min- imize^their^cytotoxicity.^For^instance,^the^non-bridging^oxygen^atoms^in^the^phosphate^group^ 28^ ^ of^the^nucleotides^forming^the^ASO^may^be^replaced^by^phosphorothioate^(PS),^resulting^in^ the^formation^of^PS^bonds^that^are^resistant^to^nuclease-based^degradation.^Moreover,^the^ 2′^position^of^the^ribose^may^be^modified^with^an^alkyl^moiety,^such^as^a^methyl^(2′-OMe)^or^ a^methoxyethyl^(2′-MOE)^group.^The^ASO^may^also^be^a^locked^nucleic^acid^(LNA),^a^peptide^ nucleic^acid^ (PNA)^ or^a^phosphorodiamidate^morpholino^ oligomer^ (PMO).^LNAs^contain^a^ constrained^ribose^ring^having^a^O2′-C4′-methylene^linkage.^PNAs^have^a^peptide-like^N-(2- aminoethyl)glycine^linkage^to^replace^the^ribose-phosphate^DNA^backbone.^PMOs^contain^a^ backbone^of^morpholine^ rings^connected^by^phosphorodiamidate^ linkages.^ Such^modified^ ASOs^are^characterized^by^a^high^target^affinity,^improved^pharmacokinetic^profiles^and^nu- clease^resistance^(Tarn^et^al,^2021).^ In^further^embodiments,^the^herein^disclosed^ASOs^may^be^gapmers,^i.e.,^chimeric^ASOs^that^ consist^of^a^central^short^region^of^deoxyribonucleotides^flanked^by^a^stretch^of^ribonucleo- tides^in^which^the^ribose^ring^is^modified^with^2′-OMe,^2′-MOE^or^LNA.^Gapmers^can^induce^ RNase^H-mediated^cleavage^of^the^target^RNA^with^a^relatively^greater^binding^affinity^and^ specificity^than^conventional^ASOs^(Tarn^et^al,^2021).^ Additional^chemical^modifications^that^can^be^introduced^into^ASOs^are^described^in^the^state^ of^the^art,^e.g.,^in^Tarn^et^al.,^2021.^Based^on^the^herein^disclosed^information^and^the^state^ of^the^art,^the^skilled^person^will^have^no^difficulties^in^designing^functional^ASOs^comprising^ the^herein^disclosed^nucleic^acid^sequences^or^subsequences^thereof.^ As^ for^ siRNAs,^ the^herein^disclosed^ASOs^may^ further^comprise^modifications^ to^ improve^ ASO^delivery.^However,^since^a^PS^backbone^assists^the^membrane^translocation^of^ASOs,^ additional^means^of^delivery^may^be^dispensable^for^PS-ASOs.^Nevertheless,^the^herein^dis- closed^ASOs^may^as^well^be^conjugated^to^suitable^lipophilic^moieties^or^anchors^such^as,^ e.g.,^ palmityl-^ (also^ referred^ to^ as^ 2′-O-hexadecyl^ or^ C16)^ or^ cholesterol-residues^ as^ de- scribed^above.^ASOs^may^also^be^conjugated^ to^cell-penetrating^peptides^ (CPPs)^ for^en- hanced^drug^delivery,^such^as^the^polycationic^HIV-1^Tat^peptide,^the^hydrophobic^residue- containing^peptide^and^artificial^poly-arginine^peptides.^ASOs^made^of^PMO^have^also^been^ conjugated^in^the^past^to^a^synthetic^octa-guanidine^dendrimer.^Such^conjugates^are^referred^ to^as^vivo-PMOs^(Tarn^et^al.,^2021).^ The^present^ invention^further^provides^an^oligonucleotide-based^therapeutic^ for^treating^an^ infection^of^HPIV,^wherein^ the^ therapeutic^comprises^one^or^more^of^ the^herein^disclosed^ oligonucleotide^ molecules^ of^ the^ invention,^ or,^ in^ case^ the^ oligonucleotide^ is^ an^ shRNA / shRNamiR,^a^nucleic^acid^encoding^them.^Suitable^delivery^methods^are^known^in^ the^art^or^described^herein.^Preferably,^the^therapeutic^comprises^^ a)^ a^lentiviral^vector^comprising^a^nucleic^acid^encoding^one^or^more^of^an^RNA^ molecule^comprising^a^sequence^having^at^least^90%^sequence^identity^to^SEQ^ ID^NO:^1^to^56,^ 29^ ^ b)^ a^lipid^nanoparticle^vector^comprising^one^or^more^of^the^herein^disclosed^oli- gonucleotide^molecules,^^ c)^ a^cationic^polymer,^selected^from^the^group^comprising^a^PEI^vector,^associ- ated^with^one^or^more^of^the^oligonucleotide^molecules^disclosed^herein,^or^ d)^ one^or^more^of^the^herein^disclosed^oligonucleotides^linked^to^a^lipophilic^moi- ety.^ The^lentiviral^vector^of^ therapeutic^a)^may^exhibit^any^of^ the^features^disclosed^herein.^As^ described^herein,^if^the^therapeutic^comprises^a^lentiviral^vector,^the^oligonucleotide^molecule^ is^ preferably^ provided^ by^ the^ vector^ as^ an^ shRNA^ or^ an^ shRNAmiR,^ preferably^ as^ an^ shRNAmiR.^ Accordingly,^ the^ lentiviral^ vector^ comprises^ a^ cargo^ sequence^ encoding^ an^ shRNA^or^an^shRNAmiR^comprising^a^nucleic^acid^sequence^having^at^least^90%,^at^ least^ 95%^or^100%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^56,^preferably^to^any^of^SEQ^ID^ NO:^1,^2,^3,^9,^12,^14,^15,^17,^18,^22,^23^or^29,^and^most^preferably^to^any^of^SEQ^ID^NO:^1^ or^23.^The^ lentiviral^vector^may^also^comprise^a^cargo^sequence^encoding^more^ than^one^ candidate^oligonucleotide^molecule^comprising^a^nucleic^acid^sequence^having^at^least^90%,^ at^least^95%^or^100%^sequence^identity^to^any^of^SEQ^ID^NO:^1^to^56.^For^instance,^it^may^ comprise^a^cargo^sequence^that^encodes^a^first^shRNA^or^an^shRNAmiR^comprising^SEQ^ID^ NO.^1^and^a^second^shRNA^or^an^shRNAmiR^comprising^SEQ^ID^NO.^23.^ Lipid^nanoparticles^for^use^as^drug^delivery^vehicles^for^siRNAs^were^first^approved^in^2018.^ A^ lipid^nanoparticle^ is^ typically^ spherical^with^ an^average^diameter^between^10^and^1000^ nanometers.^Solid^lipid^nanoparticles^possess^a^solid^lipid^core^matrix^that^can^solubilize^lip- ophilic^molecules.^The^lipid^core^is^typically^stabilized^by^surfactants^(emulsifiers).^The^emul- sifier^used^depends^on^administration^routes^and^is^more^limited^ for^parenteral^administra- tions.^The^ lipids^used^for^generating^a^ lipid^nanoparticle^may^include^ triglycerides^(e.g.^ tri- stearin),^diglycerides^(e.g.^glycerol^bahenate),^monoglycerides^(e.g.^glycerol^monostearate),^ fatty^ acids^ (e.g.^ stearic^ acid),^ steroids^ (e.g.^ cholesterol),^ and^waxes^ (e.g.^ cetyl^ palmitate)^ (https: / / en.wikipedia.org / wiki / Solid_lipid_nanoparticle).^Typical^next^generation^lipid^nanopar- ticle^compositions^also^comprise^varying^composition^of^structural^lipids^(e.g.^MC3),^choles- terol^or^beta-sitosterol,^PEG^(e.g.,^DSPG-PEG2000^or^DMG-PEG2000)^and^ionisable^or^cat- ionic^lipids^(e.g.,^DOPE^or^DOTAP).^^^ The^oligonucleotide^molecules^of^the^invention^may^also^be^delivered^in^association^with^cat- ionic^polymers^such^as,^e.g.,^polyethylenimine^(PEI)^or^DEAE-dextran,^or^branched^polymeric^ molecules,^e.g.,^hyperbranched^polymers^such^as^hyperbranched^polyglycerol^polymers^or^ dendrimers.^The^association^can^be^covalent^or^non-covalent,^but^typically^is^non-covalent.^ The^cationic^polymer,^ e.g.,^polyethyleneimine^ (PEI)^ is^ capable^of^complexing^with^nucleic^ acids^via^electrostatic^ interactions^between^ its^cationic^groups^and^ the^negatively^charged^ nucleic^acids,^thereby^forming^so^called^polyplexes.^These^polyplexes^are^characterized^by^ 30^ ^ high^transfection^efficiency^due^to^the^high^buffering^capacity^of^PEI,^which^facilitates^endo- somal^escape^of^the^gene^payload^(Schwarz^and^Merkel^et^al.,^2017).^The^cationic^polymers,^ e.g.,^the^PEI^vectors^can^be^functionalized^with^a^wide^variety^of^polymers^including^oligosac- charides,^ poly(ethylene^glycol)^ (PEG),^poly(ε-caprolactone),^ and^others.^ They^may^ further^ carry,^e.g.,^targeting^moieties^or^comprise^chemical^modifications^such^as^ester^or^disulfide^ bonds^to^yield^degradable^PEI^analogs^that^exhibit^reduced^cytotoxicity.^^ As^already^described^herein,^the^delivery^of^the^oligonucleotide^candidate^molecule^may^also^ be^improved^by^directly^linking^the^oligonucleotide^molecule^to^lipophilic^moieties.^Exemplary^ lipophilic^moieties^for^delivering^RNAi^triggers^such^as^siRNAs^or^for^delivery^of^ASOs^include^ lipid^residues^such^as^palmityl-^ (also^ referred^ to^as^2′-O-hexadecyl^or^C16)^or^cholesterol- residues^as^described,^e.g.,^in^Raouane^et^al.,^2012^or^Brown^et^al.,^2022.^Especially^2′-O- hexadecyl^(C16)^conjugates^facilitate^enhanced^delivery^and^siRNA^uptake^into^the^alveolar^ and^bronchiolar^epithelium,^which^is^why^they^may^be^particularly^preferred^for^delivering^the^ herein^disclosed^candidate^oligonucleotide^molecules^to^cells^of^the^respiratory^tract,^e.g.,^via^ inhalative^administration.^ The^inventors^found^that,^if^the^oligonucleotide^molecules^disclosed^herein^are^provided^as^ siRNA^consisiting^of^an^antisense^(guide^strand)^and^a^sense^strand^(passenger^strand)^com- plementary^to^the^antisense^sense,^it^is^advantageous^to^conjugate^the^palmityl-^(C16-)^resi- due^either^to^ the^3’^end,^the^5’^end^or^at^position^6^(N6)^of^the^sense^strand^of^ the^siRNA^ doublex.^As^shown^in^the^examples^below,^linking^a^palmityl-residue^at^one^of^these^specific^ positions^within^the^sense^strand^of^the^herein^disclosed^siRNAs^and^delivering^the^thus^mod- ified^siRNAs^in^a^parainfluenza^infection^experiment^to^infected^LLCMK2^cells^or^human^lung^ slices^resulted^in^a^reduction^in^viral^load^that^was^comparable^to^the^reduction^achieved^when^ the^siRNAs^were^administered^using^nebulized^LNPs^or^lipofectamine.^ Thus,^in^a^preferred^embodiment,^the^oligonucleotide-based^therapeutic^of^the^invention^com- prises^one^or^more^of^the^herein^disclosed^synthetic^siRNA^consisting^of^an^antisense^and^a^ complementary^sense^strand,^wherein^the^sense^strand^comprises^a^palmityl-residue^(i.e.,^a^ 2′-O-hexadecyl)^ conjugated^ to^ its^3’^ end,^ its^5’^ end^and / or^ to^ the^nucleotide^ at^nucleotide^ position^6.^In^the^context^of^the^invention,^the^term^“nucleotide^position^6”^refers^to^the^sixth^ nucleotide^starting^from^the^5'^end^of^the^passenger / sense^strand.^ For^ instance,^disclosed^herein^is^an^oligonucleotide-based^therapeutic^comprising^a^siRNA^ consisting^of^an^antisense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^169^and^a^ complementary^sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^170,^wherein^ the^sense^strand^comprises^a^palmityl-residue^conjugated^to^its^3’^end.^ Alternatively,^the^oligonucleotide-based^therapeutic^comprises^a^siRNA^consisting^of^an^an- tisense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^169^and^a^complementary^ 31^ ^ sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^170,^wherein^the^sense^strand^ comprises^a^palmityl-residue^conjugated^to^its^5’^end.^ In^yet^another^embodiment,^the^oligonucleotide-based^therapeutic^comprises^a^siRNA^con- sisting^of^ an^antisense^strand^having^a^nucleic^ acid^sequence^of^SEQ^ ID^NO:^ 169^and^a^ complementary^sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^170,^wherein^ the^sense^strand^comprises^a^palmityl-residue^conjugated^to^the^nucleotide^at^nucleotide^po- sition^6^o^SEQ^ID^NO:^170.^ Also^disclosed^herein^is^an^oligonucleotide-based^therapeutic^comprising^a^siRNA^consisting^ of^an^antisense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^173^and^a^comple- mentary^sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^174,^wherein^the^sense^ strand^comprises^a^palmityl-residue^conjugated^to^its^3’^end.^ Alternatively,^the^oligonucleotide-based^therapeutic^comprises^a^siRNA^consisting^of^an^an- tisense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^173^and^a^complementary^ sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^174,^wherein^the^sense^strand^ comprises^a^palmityl-residue^conjugated^to^its^5’^end.^ In^yet^another^embodiment,^the^oligonucleotide-based^therapeutic^comprises^a^siRNA^con- sisting^of^ an^antisense^strand^having^a^nucleic^ acid^sequence^of^SEQ^ ID^NO:^ 173^and^a^ complementary^sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^174,^wherein^ the^sense^strand^comprises^a^palmityl-residue^conjugated^to^the^nucleotide^at^nucleotide^po- sition^6^o^SEQ^ID^NO:^174.^ Also^disclosed^herein^is^an^oligonucleotide-based^therapeutic^comprising^a^siRNA^consisting^ of^an^antisense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^175^and^a^comple- mentary^sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^176,^wherein^the^sense^ strand^comprises^a^palmityl-residue^conjugated^to^its^3’^end.^ Alternatively,^the^oligonucleotide-based^therapeutic^comprises^a^siRNA^consisting^of^an^an- tisense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^175^and^a^complementary^ sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^176,^wherein^the^sense^strand^ comprises^a^palmityl-residue^conjugated^to^its^5’^end.^ In^yet^another^embodiment,^the^oligonucleotide-based^therapeutic^comprises^a^siRNA^con- sisting^of^ an^antisense^strand^having^a^nucleic^ acid^sequence^of^SEQ^ ID^NO:^ 175^and^a^ complementary^sense^strand^having^a^nucleic^acid^sequence^of^SEQ^ID^NO:^176,^wherein^ the^sense^strand^comprises^a^palmityl-residue^conjugated^to^the^nucleotide^at^nucleotide^po- sition^6^o^SEQ^ID^NO:^176.^ As^described^herein,^when^the^vector^ is^a^ lipid^nanoparticle,^a^cationinc^polymer^such^as,^ e.g.,^a^PEI^vector^or^a^lipophilic^moiety^such^as^2′-O-hexadecyl,^the^oligonucleotide^molecule^ 32^ ^ is^preferably^provided^in^the^form^of^any^of^the^herein^disclosed^siRNAs^or^ASOs.^The^oligo- nucleotide^ molecule^ may^ however^ also^ be^ provided^ as^ an^ in^ vitro^ produced^ shRNA^ or^ shRNAmiR.^In^some^embodiments,^ the^ lipid^nanoparticle^or^the^cationic^polymer,^e.g.,^the^ PEI^vector^comprises^only^oligonucleotides^molecules^comprising^or^consisting^of^the^same^ nucleic^acid,^e.g.,^the^lipid^nanoparticle^comprises^only^RNA^molecules^comprising^or^con- sisting^of^a^nucleic^acid^sequence^of^SEQ^ID^NO:^1^or^ASOs^comprising^or^consisting^of^SEQ^ ID^NO:^204.^Alternatively,^the^lipid^nanoparticle^or^the^cationic^polymer,^e.g.,^the^PEI^vector,^ may^also^comprise^a^mixture^of^oligonucleotide^molecules^comprising^or^consisting^of^differ- ent^nucleic^acid^sequences,^e.g.^RNA^molecules^comprising^or^consisting^of^SEQ^ID^NO:^1^ and^RNA^molecules^comprising^or^consisting^of^SEQ^ID^NO:^18,^RNA^molecules^comprising^ or^consisting^of^SEQ^ID^NO:^1^and^RNA^molecules^comprising^or^consisting^of^SEQ^ID^NO:^ 23^or^RNA^molecules^comprising^or^consisting^of^SEQ^ID^NO:^18^and^RNA^molecules^com- prising^or^consisting^of^SEQ^ID^NO:^23.^Alternatively,^the^lipid^nanoparticle^or^ the^cationic^ polymer,^e.g.,^the^PEI^vector,^may^also^comprise^a^mixture^of^different^ASOs^disclosed^herein,^ e.g.,^ASOs^of^SEQ^ID^NO:^204^and^SEQ^ID^NO:^205,^ASOs^of^SEQ^ID^NO:^205^and^206,^or^ ASOs^of^SEQ^ID^NO:^204,^205^and^206.^The^lipid^nanoparticle^or^the^cationic^polymer,^e.g.,^ the^PEI^vector,^may^also^comprise^a^mixture^of^any^of^the^herein^disclosed^RNAi^triggers^and^ ASOs,^e.g.,^it^may^comprise^RNA^molecules^comprising^or^consisting^of^SEQ^ID^NO:^1^and^ ASOs^consisting^of^SEQ^ ID^NO:^ 204.^ In^ such^a^ scenario,^ the^RNA^molecules^and^ASOs^ present^in^the^lipid^nanoparticle^or^cationic^polymer^may^bind^to^the^same^target^sequence^or^ to^different^ones.^ The^therapeutic^may^also^comprise^several^lipid^nanoparticles,^cationic^polymers^such^as^PEI^ vectors^or^oligonucleotide-conjugates^as^described^herein.^In^case^the^therapeutic^comprises^ several^lipid^nanoparticles^or^cationic^polymers^such^as^PEI^vectors,^each^lipid^nanoparticle^ or^cationic^polymer^comprises^an^oligonucleotide^molecule^comprising^or^consisting^of^a^dif- ferent^nucleic^acid^sequence^disclosed^herein.^For^instance,^the^therapeutic^may^comprise^a^ first^ lipid^nanoparticle^and / or^cationic^polymer,^comprising^an^RNA^molecule^comprising^or^ consisting^of^a^nucleic^acid^sequence^of^SEQ^ID^NO:^1,^a^second^lipid^nanoparticle^and / or^ cationic^polymer^comprising^an^RNA^molecule^comprising^or^consisting^of^a^nucleic^acid^se- quence^of^SEQ^ ID^NO:^18,^and,^optionally,^even^a^ third^ lipid^nanoparticle^and / or^ cationic^ polymer^comprising^an^RNA^molecule^comprising^or^consisting^of^a^nucleic^acid^sequence^of^ SEQ^ID^NO:^23.^The^ therapeutic^may^also^comprise,^e.g.,^a^ first^ lipid^nanoparticle^and / or^ cationic^polymer,^comprising^an^ASO^of^SEQ^ID^NO:^204,^a^second^lipid^nanoparticle^and / or^ cationic^polymer^comprising^an^ASO^of^SEQ^ID^NO:^205,^and,^optionally,^even^a^third^lipid^ nanoparticle^and / or^cationic^polymer^comprising^an^ASO^of^SEQ^ID^NO:^206.^The^therapeutic^ may^ also^ comprise^ several^ lipid^ nanoparticles^ or^ cationic^ polymers^ such^ as^ PEI^ vectors,^ wherein^some^lipid^nanoparticle^or^cationic^polymers^within^the^therapeutic^comprise^any^of^ the^herein^disclosed^RNA^molecules^and^some^lipid^nanoparticle^or^cationic^polymers^within^ the^therapeutic^comprise^any^of^the^herein^disclosed^ASOs.^For^instance,^the^therapeutic^may^ 33^ ^ comprise^a^first^lipid^nanoparticle^and / or^cationic^polymer,^comprising^an^RNA^molecule^com- prising^or^consisting^of^a^nucleic^acid^sequence^of^SEQ^ID^NO:^1,^a^second^lipid^nanoparticle^ and / or^cationic^polymer^comprising^an^ASO^of^SEQ^ID^NO:^205.^In^such^a^scenario,^the^RNA^ molecules^and^ASOs^used^in^the^therapeutic^may^either^bind^to^the^same^target^sequence^or^ to^different^ones,^preferably^ to^different^ones.^The^ therapeutic^may^also^comprise^several^ different^oligonucleotide^molecules^of^the^invention^linked^to^a^liphophilic^moiety^such^as^2′- O-hexadecyl^(C16).^For^instance,^the^therapeutic^may^comprise^a^mixture^of^RNA^molecules^ comprising^ or^ consisting^of^ SEQ^ ID^NO:^ 1^ conjugated^ to^ 2′-O-hexadecyl,^RNA^molecules^ comprising^or^consisting^of^SEQ^ID^NO:^18^conjugated^to^2′-O-hexadecyl,^and / or^RNA^mole- cules^comprising^or^consisting^of^SEQ^ID^NO:^23^conjugated^to^2′-O-hexadecyl.^^ In^yet^another^aspect,^the^present^invention^further^provides^a^composition^for^use^in^treating^ a^respiratory^disease^in^a^subject,^wherein^said^composition^is^administered^to^the^airways^of^ the^subject.^The^composition^comprises^a^lentiviral^vector^pseudotyped^with^a^vesiculoviral^ envelope^glycoprotein^and^at^least^one^surface^protein^of^influenza^A^virus^and / or^at^least^one^ surface^protein^of^respiratory^syncytial^virus.^^ In^the^context^of^ the^invention,^a^respiratory^disease^may^be^any^pathological^condition^af- fecting^the^respiratory^tract^including^the^trachea,^bronchi,^bronchioles,^alveoli,^pleurae,^pleu- ral^cavity,^the^nerves^and^muscles^of^respiration.^Exemplary^respiratory^diseases^may^be^the^ common^cold,^ influenza,^COVID-19^or^pharyngitis,^but^may^also^ include^more^severe^dis- eases^such^as^lung^cancer.^ The^composition^may^be^administered^to^the^airways^of^a^subject^by^any^suitable^means^and^ by^any^suitable^route^known^to^the^skilled^person.^For^instance,^the^composition^may^be^in- jected^into^the^bloodstream^of^the^subject.^In^a^preferred^embodiment,^the^composition^how- ever^reaches^the^airways^of^the^subject^via^inhalation,^i.e.,^the^composition^may^be^provided^ to^the^patient^as^an^aerosol.^In^the^context^oft^the^invention,^the^term^„airways“^refers^to^the^ organs^of^ the^ respiratory^ tract^of^a^subject.^The^airways^can^be^subdivided^ into^ the^upper^ airways^and^lower^airways.^The^upper^airways^comprise^the^trachea^as^well^as^the^main^and^ lobar^bronchi^,^whereas^the^lower^airways^comprise^the^smaller^conducting^bronchi^and^bron- chioles.^ A^ lentiviral^vector^ is,^as^ the^name^ implies,^derived^ from^a^genus^of^ retroviruses^known^as^ lentivirus.^Lentiviruses^comprise^e.g.^human^immunodeficiency^virus^(HIV),^simian^immuno- deficiency^virus^(SIV),^feline^immunodeficiency^virus^(FIV)^or^equine^infectious^anemia^virus^ (EIAV).^ Like^ that^ of^ other^ retroviruses,^ the^ genome^ of^ lentiviruses^ consists^ of^ a^ single- stranded^(ss)^positive^sense^RNA.^During^replication,^the^ lentiviral^ssRNA^genome^ is^con- verted^into^double-stranded^(ds)^DNA^by^a^process^known^as^reverse^transcription.^The^re- verse^ transcribed^ lentiviral^dsDNA^ is^subsequently^ integrated^ into^ the^host^ cell’s^genome,^ 34^ ^ which^ in^ turn^ replicates^ and^ transcribes^ the^ integrated^ lentiviral^ genes^ along^with^ its^ own^ genes^to^produce^new^viral^particles.^^ The^lentiviral^vector^in^the^herein^disclosed^composition^may^thus^be^derived^from^a^lentivirus^ selected^from^the^group^comprising^HIV-1,^HIV-2,^SIV,^FIV^or^EIAV.^In^a^preferred^embodi- ment,^the^lentiviral^vector^is^derived^from^HIV-1.^^ The^lentiviral,^e.g.,^HIV-1,^genome^comprises^three^major^structural^genes:^gag,^pol^and^env.^ gag^encodes^ the^viral^matrix^ (MA),^capsid^(CA)^and^nucleocapsid^ (NC),^which^collectively^ facilitate^the^assembly^and^release^of^the^virus^particles.^The^pol^gene^encodes^the^viral^en- zymes^protease^(PR),^reverse^transcriptase^(RT)^and^integrase^(IN),^which^govern^viral^rep- lication.^The^HIV-1^env^encodes^the^viral^surface^glycoprotein^gp160,^which^is^subsequently^ cleaved^to^form^the^surface^protein^gp120^and^the^transmembrane^protein^gp41^during^viral^ maturation.^In^addition^to^the^structural^genes^gag,^pol^and^env,^the^HIV-1^genome^further- more^comprises^the^two^regulatory^genes^tat^and^rev^as^well^as^the^four^accessory^genes^vif,^ vpr,^vpu^and^nef.^Whereas^ tat^encodes^a^transactivator^required^ for^viral^ transcription,^ rev^ encodes^a^protein^that^controls^both^splicing^and^export^of^viral^transcripts.^The^four^acces- sory^genes^are^considered^non-essential^for^viral^replication,^but^are^believed^to^increase^its^ efficiency^(German^Advisory^Committee^Blood^(Arbeitskreis^Blut),^Subgroup^‘Assessment^of^ Pathogens^Transmissible^by^Blood’,^2016,^Human^Immunodeficiency^Virus^(HIV).^Transfus^ Med^Hemother.^43(3),^203-222).^ The^lentiviral^vector^used^in^the^herein^disclosed^composition^preferably^is^a^third-generation^ lentiviral^vector.^ In^1996,^Naldini^et^al.^reported^the^use^of^an^HIV-based^vector^for^in^vivo^gene^delivery^and^ stable^transduction^of^non-dividing^cells^(Naldini^et^al.,^1996).^Use^of^HIV-1-derived^lentiviral^ vectors^for^in^vivo^gene^transfer^was^however^associated^with^considerable^health^risks.^To^ meet^these^biosafety^concerns,^the^first^generation^of^lentiviral^vector^systems^split^the^viral^ genome^into^three^separate^plasmids^to^avoid^the^formation^of^replication-competent^viruses.^ The^first^plasmid^encoded^the^actual^vector^that^was^to^be^integrated^into^the^host^cell’s^ge- nome.^It^comprised^ the^ transgene^of^ interest^ functionally^ linked^ to^a^suitable^promoter^se- quence^as^well^as^cis-elements^necessary^ for^polyadenylation,^ integration,^ initiation^of^ re- verse^transcription^and^packaging^(e.g.,^the^long-terminal^repeats,^the^packaging^signal,^the^ primer^binding^site,^the^polypurine^tract^or^the^Rev-responsive^element).^The^second^plasmid,^ known^as^packaging^plasmid,^comprised^the^genes^encoding^the^viral^proteins^that^contribute^ to^ packaging,^ reverse^ transcription^and^ integration^of^ the^ viral^ genome,^ i.e.,^gag,^pol,^ the^ regulatory^genes^tat^and^rev^as^well^as^the^four^accessory^genes^vif,^vpu,^vpr^and^nef.^The^ third^plasmid^(Env^plasmid)^expressed^ the^viral^glycoprotein^for^host^cell^receptor^binding.^ Due^to^the^physical^separation^of^the^packaging^genes^from^the^rest^of^the^viral^genome,^this^ split^genome^design^prevented^viral^replication^after^infection^of^the^host^cell.^ 35^ ^ To^further^ improve^the^safety^of^lentiviral^vectors,^a^second^generation^system^was^estab- lished^by^removing^all^accessory^genes^from^the^packaging^plasmid,^as^they^were^found^to^ constitute^crucial^virulence^factors.^ With^the^third^generation^of^lentiviral^vector^systems,^so-called^self-inactivating^(SIN)^vectors^ were^introduced.^When^lentiviral^vectors^are^integrated^into^a^host^cell^genome,^the^transgene^ cassette^of^the^provirus^comprising^the^gene^of^interest^is^flanked^by^two^long^terminal^repeats^ (LTRs).^The^presence^of^these^LTRs^may^promote^the^emergence^of^potentially^harmful^rep- lication-competent^recombinants.^In^addition,^viral^promoter / enhancer^regions^located^in^the^ LTRs^could^induce^the^expression^of^adjacent^host^genes,^with^potentially^tumorigenic^con- sequences.^Moreover,^the^promoter / enhancer^regions^in^these^LTRs^can^transcriptionally^in- terfere^with^the^promoter^driving^the^transgene^as^well^as^the^neighboring^genes.^Therefore,^ promoter / enhancer^sequences^of^the^viral^3’^LTR^were^removed^by^deleting^a^particular^re- gion^inside^the^LTR^(U3)^from^the^DNA^that^was^used^to^produce^the^viral^RNA.^Deletion^of^ the^U3^region^effectively^abolished^the^transcriptional^activity^of^the^LTR.^Furthermore,^tat,^a^ regulatory^gene^driving^viral^transcription,^was^deleted^from^the^packaging^plasmid,^whereas^ the^second^regulatory^gene^rev^was^provided^from^another^separate,^fourth,^plasmid^(Zufferey^ et^al.,^1998;^Schambach^et^al.,^2013).^In^case^the^5’^promoter^driving^the^viral^genomic^mRNA^ is^not^HIV-derived^and^substituted^by^a^CMV^or^RSV^promoter,^an^extra^tat^plasmid^may^be^ omitted.^ Accordingly,^ the^ lentiviral^vector^used^in^the^herein^disclosed^composition^ is^ replication^ in- competent^due^to^the^split^packaging^design^and^self-inactivating^(SIN)^due^to^a^deletion^in^ the^U3^region^of^the^3’^LTR.^It^further^lacks^the^genes^vif,^vpr,^vpu,^nef,^and,^optionally,^tat.^^ Therefore,^the^lentiviral^vector^in^the^herein^disclosed^composition^preferably^comprises^the^ following^features:^ a)^ a^5’^LTR^comprising^a^constitutively^active^heterologous^promoter^at^the^U3^position,^a^ repeat^region^(R)^and^a^U5^region,^ b)^ a^5’^UTR^comprising^a^primer^binding^site^(PBS),^a^splice^donor^site^(SD),^a^packaging^ signal^(ψ),^a^Rev-responsive^element,^and,^optionally,^a^splice^acceptor^(SA)^site,^ c)^ an^internal^enhancer / promoter^region^operably^linked^to^a^cargo^sequence,^ d)^ RNA^processing^elements^optionally^comprising^a^Woodchuck^hepatitis^virus^posttran- scriptional^regulatory^element^(PRE),^and^^ e)^ ^a^3’^LTR^with^a^deleted^(SIN)^U3^region,^a^repeat^region^(R)^and^a^U5^region.^ The^splice^acceptor^site^is^optional.^The^splice^donor^site^is^important,^but^vectors^lacking^it^ are^functional.^Presence^of^RNA^processing^elements^such^as^a^Woodchuck^hepatitis^virus^ posttranscriptional^ regulatory^ element^ (PRE)^ contribute^ to^ efficiency,^ and^ vectors^ without^ them^are^typically^not^generated^in^high^titers.^However,^they^can^be^concentrated.^ 36^ ^ Suitable^3rd^generation^lentiviral^vectors^are^e.g.,^known^in^the^art^and / or^can^be^prepared^by^ the^skilled^person.^An^exemplary^lentiviral^vector^that^can^be^used^in^the^invention^comprises^ a^nucleic^acid^sequence^of^SEQ^ID^NO:^179.^The^lentiviral^vector^may^also^comprise^a^nucleic^ acid^sequence^having^at^least^80%,^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^ at^least^97%,^at^least^98%,^or^at^least^99%^sequence^identity^to^SEQ^ID^NO:^179.^It^may^also^ consist^of^SEQ^ID^NO:^179.^Alternatively,^the^lentiviral^vector^according^to^the^invention^may^ comprise^a^different^promoter^region^than^SEQ^ID^NO:179,^e.g.,^another^suitable^promotor^ as^taught^herein.^ Optionally,^the^viral^enhancer / promoter^regions^located^at^the^U3^position^of^the^5’^LTR^in^the^ lentiviral^vector^may^be^replaced^by^a^heterologous^promoter^to^enhance^expression^of^ge- nomic^RNA^and^to^compensate^for^the^loss^of^the^lentiviral^Tat^protein.^Therefore,^the^heter- ologous^promoter^preferably^ is^a^ strong^promoter,^ i.e.,^ it^ drives^constitutive^ expression^ of^ genes^under^its^control.^It^may^be^selected^from^the^group^comprising^a^Rous^sarcoma^virus^ (RSV)^promoter,^a^human^cytomegalovirus^(CMV)^promoter,^an^HIV^promoter^(in^case^a^len- tiviral^vector^is^used^that^is^not^derived^from^HIV-1^or^HIV-2)^or^a^eukaryotic^promoter^such^ as^e.g.,^EF1a,^PGK1,^UBC,^or^human^beta^actin.^It^may^also^be^an^inducible^promoter^that^ only^drives^gene^expression^in^the^presence^or^absence^of^a^certain^stimulus,^in^particular,^ when^expression^of^the^vector^would^be^toxic^for^the^packaging^cell.^It^may,^e.g.,^be^a^Tet- regulated^promoter,^i.e.,^it^may^comprise^a^tetracycline^response^element^(TRE)^that^can^be^ bound^by^a^tetracycline^transactivator^(tTA)^protein^in^the^presence^of^tetracycline^or^an^ana- logue^thereof,^e.g.^doxycycline.^Preferably,^the^heterologous^promoter^is^a^constitutively^ac- tive^promoter^derived^from^RSV^or^CMV.^ The^5’^UTR^packaging^signal^(ψ)^interacts^with^the^nucleocapsid^(NC)^of^the^assembling^Gag^ proteins^to^mediate^packaging.^Optimal^packaging^furthermore^requires^the^trans-activation^ response^element^(TAR),^U5,^ the^primer^binding^site^(PBS)^and^the^group-specific^antigen^ (Gag).^The^Rev^protein^additionally^enhances^the^encapsidation^of^RNA^containing^the^Rev- responsive^element^(RRE).^The^RRE^also^allows^for^higher^expression^of^the^viral^genome^ and^may^suppress^aberrant^splice^events^(Schambach^et^al.,^2013).^The^5’^UTR^may,^option- ally,^further^comprise^multiple^splice^sites.^In^particular,^the^lentiviral^vector^may^comprise^a^ 5’UTR^splice^donor^(SD)^site^and,^optionally,^a^splice^acceptor^site^(SA).^In^an^optional^em- bodiment,^the^5’^UTR^may^further^comprise^a^polypurine^tract^(PPT)^region,^preferably^down- stream^of^the^RRE.^The^PPT^region^is^thought^to^increase^the^efficiency^of^reverse^transcrip- tion^and^may^even^mediate^nuclear^entry^of^the^lentiviral^pre-integration^complex,^which^forms^ after^uncoating^of^the^viral^particle^upon^entry^into^the^host^cell^(Schambach^et^al.,^2013).^ The^internal^promoter / enhancer^region^is^preferably^a^strong^promoter,^i.e.,^it^drives^constitu- tive^expression^of^the^cargo^sequence^under^its^control.^It^may^be^a^promoter^selected^from^ the^group^comprising^a^viral^promoter,^a^cellular^promoter,^a^cell-specific^promoter,^an^induc- ible^promoter^or^a^synthetic^promoter.^^ 37^ ^ It^may,^e.g.,^be^a^viral^promoter^selected^from^the^group^comprising^a^promoter^derived^from^ CMV,^spleen^focus-forming^virus^(SFFV),^myeloproliferative^sarcoma^virus^(MPSV),^murine^ embryonal^stem^cell^virus^(MESV),^murine^leukemia^virus^(MLV)^and^simian^virus^40^(SV40).^ Preferably,^the^internal^promoter^is^a^promoter^derived^from^CMV^or^SFFV,^most^preferably^ from^CMV.^In^another^preferred^embodiment,^the^promoter^may^also^be^a^CAG^promoter,^i.e.^ a^composite^construct^consisting^of^the^CMV^enhancer^fused^to^the^chicken^beta-actin^pro- moter^and^the^rabbit^beta-Globin^splice^acceptor^site.^The^internal^promoter / enhancer^region^ may^ alternatively^ be^ a^ cellular^ promoter^ selected^ from^ the^ group^ comprising^EF1a,^ EFS,^ PGK1^and^UBC.^ The^internal^promoter / enhancer^may^also^be^an^inducible^promoter,^especially,^if^expression^ of^the^cargo^sequence^may^lead^to^cytotoxicity.^The^promoter^may^thus^be^selected^from^the^ group^comprising^a^TET-inducible^promoter^or^a^Cumate-inducible^promoter.^ The^internal^promoter / enhancer^region^may^also^be^a^hybrid^or^a^synthetic^promoter^selected^ from^the^group^comprising^CAG^and^MND.^A^hybrid^promoter^is^composed^of^several^regu- latory^elements^from^different^promoters / enhancers,^which^are^joined^to^build^a^new^promoter^ combining^the^desired^features.^^In^contrast,^a^synthetic^promoter^essentially^consists^of^an^ array^ of^ transcription^ factor^binding^sites^ fused^ to^ a^minimal^ promoter.^A^preferred^hybrid^ promoter^used^for^driving^cargo^sequence^expression^is^CAG.^ Finally,^the^internal^enhancer / promoter^region^may^comprise^a^chromatin^opening^element,^ i.e.,^a^DNA^sequence^consisting^of^methylation-free^CpG^islands^that^either^encompasses^a^ housekeeping^gene^promoter^or^is^operably^linked^to^a^heterologous^promoter^to^confer^re- producible,^stable^transgene^expression.^ The^3’^UTR^of^ the^ lentiviral^ vector^according^ to^ the^ invention^may^comprise^an^RNA^pro- cessing^element,^ e.g.,^ a^Woodchuck^ hepatitis^virus^ posttranscriptional^ regulatory^element^ (PRE).^The^PRE^may^enhance^cargo^sequence^expression^by^up^to^10-fold^and^contributes^ to^high^virus^titers^(Schambach^et^al.,^2013).^^ To^further^prevent^read-through^transcription,^the^3’^end^of^the^lentiviral^vector^may^comprise^ upstream^sequence^elements^(USE)^that^can^be^inserted^into^the^3’^U3^region^to^improve^3’^ end^processing^and^transcriptional^termination.^Preferably,^the^USE^is^a^recombinant^direct^ repeat^of^the^USE^derived^from^SV40^(2xSV^USE).^The^3’^UTR^may,^optionally,^further^com- prise^target^sites^for^differentiation-specific^cellular^miRNAs^to^enable^post-transcriptional^tar- geting^of^the^vector^mRNA.^^ Upon^random^or^semi-random^ integration^of^a^ lentiviral^vector^ into^or^next^ to^a^host^cell’s^ gene,^the^vector^may^act^as^a^mutagen^that^dysregulates^the^gene’s^expression.^^In^an^op- tional^embodiment,^the^space^created^by^deletion^of^the^U3^region^in^the^3’LTR^of^the^lentiviral^ vector^may^therefore^comprise^an^insulator^sequence.^Insulator^sequences^constitute^barriers^ 38^ ^ between^regulator^gene^regions^and^thus^may^prevent^vector-driven^dysregulation^of^adjacent^ proto-oncogenes,^i.e.,^cellular^genes^whose^erroneous^expression^may^provoke^tumorigene- sis^(Schambach^et^al.,^2013).^ To^ further^ increase^biosafety^of^ the^composition^according^to^ the^invention,^ the^packaging^ plasmid^encoding^for^Gag-Pol^may^be^designed^to^comprise^an^inhibitory^mutation^in^the^pol^ region^encoding^for^the^integrase^protein^to^render^the^lentiviral^vector^integration-deficient.^ Possible^mutations^may^alter^a^triad^of^amino^acid^residues^at^the^catalytic^core^of^the^inte- grase^enzyme,^e.g.,^D64,^D116^and^E152.^Alternatively,^amino^acid^changes^may^be^intro- duced^into^the^integrase^DNA^attachment^site^e.g.^by^mutating^nucleotides^within^a^12^base- pair^(bp)^region^of^the^vector^5’^LTR^U3^region^or^an^11^bp^region^of^the^3’LTR^U5^region^ (Shaw^and^Cornetta,^2014).^Therefore,^ in^an^optional^embodiment,^ the^lentiviral^vector^ac- cording^to^the^invention^is^a^non-integrating^lentiviral^vector.^^ The^lentiviral^vector^is^preferably^pseudotyped^with^a^vesiculoviral^envelope^glycoprotein^and^ at^least^one^surface^protein^of^influenza^A^virus^and / or^at^least^one^surface^protein^of^respir- atory^syncytial^virus.^ Vesiculovirus^refers^to^a^genus^of^negative-sense^single-stranded^RNA^viruses^in^the^family^ Rhabdoviridae,^within^the^order^Mononegavirales.^ The^vesicular^envelope^glycoprotein^is^selected^from^the^group^comprising^MaraVg,^VSVg,^ CoCVg^and^PyriVg.^ Accordingly,^in^a^first^embodiment,^the^vesicular^envelope^glycoprotein^used^for^pseudotyping^ the^lentiviral^vector^is^MaraVg^and^thus^may^have^an^amino^acid^sequence^having^at^least^ 80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^ 98%,^at^ least^ 99%^or^100%^sequence^ identity^ to^SEQ^ ID^NO:^ 180.^Optionally,^ it^may^be^ encoded^by^a^nucleic^acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^least^ 90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^or^100%^sequence^ identity^to^SEQ^ID^NO:^181.^MaraVg^is^an^envelope^glycoprotein^derived^from^Maraba^virus.^ Maraba^virus^is^a^rhabdovirus^best^known^for^displaying^broad^tropism^for^both^human^and^ murine^cancer^cells.^^ In^another^embodiment,^the^vesicular^envelope^glycoprotein^used^for^pseudotyping^the^lenti- viral^vector^is^VSVg,^e.g.,^wild-type^VSVg^or^a^VSVg^derivative^capable^of^binding^to^the^LDL- receptor^or^LDL-R^family^members.^Preferably,^the^wild^type^VSVg^is^a^glycoprotein^derived^ from^the^Indiana^VSV^serotype^and^has^an^amino^acid^sequence^having^at^least^80%,^pref- erably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^ least^ 99%^or^ 100%^sequence^ identity^ to^SEQ^ ID^NO:^182.^Optionally,^ it^ is^encoded^by^ a^ nucleic^acid^sequence^having^at^ least^80%,^preferably^at^ least^85%,^at^ least^90%,^at^ least^ 95%,^at^ least^96%,^at^least^97%,^at^ least^98%,^at^least^99%^or^100%^sequence^ identity^to^ 39^ ^ SEQ^ID^NO:^183.^It^may^also^be^derived^from^the^New^Jersey^VSV^serotype^and^have^an^ amino^acid^sequence^having^at^ least^80%,^preferably^at^ least^85%,^at^ least^90%,^at^ least^ 95%,^at^ least^96%,^at^least^97%,^at^ least^98%,^at^least^99%^or^100%^sequence^ identity^to^ SEQ^ID^NO:^184.^Optionally,^it^is^encoded^by^a^nucleic^acid^sequence^having^at^least^80%,^ preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^ at^least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^185.^To^achieve^higher^particle^sta- bility^upon^in-vivo^administration^and^to^evade^potential^recognition^by^the^host’s^complement^ system,^a^thermostable^and^complement-resistant^VSVg^glycoprotein^(VSVg^ts)^may^alterna- tively^be^used,^which^may^have^an^amino^acid^sequence^having^at^least^80%,^preferably^at^ least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^ or^100%^sequence^identity^to^SEQ^ID^NO:^186.^It^may,^optionally,^be^encoded^by^a^nucleic^ acid^sequence^having^at^least^80%,^preferably^at^ least^85%,^at^least^90%,^at^least^95%,^at^ least^96%,^at^ least^97%,^at^least^98%,^at^least^99%^or^100%^sequence^ identity^to^SEQ^ID^ NO:^187,^and^capable^of^binding^to^the^LDL-R^or^LDL-R^ family^members.^VSVg^confers^a^ very^broad^tropism^to^viral^vectors.^Upon^binding^of^VSVg^to^the^LDL-receptor,^the^virion^is^ taken^up^by^the^host^cell^via^clathrin-mediated^endocytosis.^Besides^providing^high^tropism,^ VSVg^furthermore^significantly^increases^the^stability^of^the^lentiviral^vector,^which^allows^for^ efficient^concentration^of^viral^titers^by^centrifugation,^another^essential^feature^for^effective,^ virus-mediated^gene^transfer.^Accordingly,^pseudotyping^of^lentiviral^vectors^with^VSV-G^al- lows^for^robust^transduction^into^a^wide^range^of^different^cell^types.^^ In^a^further^embodiment,^the^vesicular^envelope^glycoprotein^used^for^pseudotyping^the^len- tiviral^vector^is^CoCVg^glycoprotein,^i.e.,^a^glycoprotein^derived^from^Cocal^virus.^CoCVg^is^ capable^of^binding^to^the^LDL-receptor^and^it^may,^e.g.,^have^an^amino^acid^sequence^having^ at^least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^ at^least^98%,^at^least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^188.^Optionally,^CoCVg^ may^be^encoded^by^a^nucleic^acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^ least^90%,^at^ least^95%,^at^ least^96%,^at^ least^97%,^at^ least^98%,^at^ least^99%^or^100%^ sequence^identity^to^SEQ^ID^NO:^189.^In^2010,^Trobridge^et^al.^addressed^some^of^the^dis- advantages^of^VSVg^-pseudotyped^lentiviral^vectors,^including^the^aforementioned^cytotoxi- city^associated^with^constitutive^VSVg^expression,^but^also^the^inactivation^of^VSVg^-pseudo- typed^vectors^by^human^serum^complement.^ The^authors^of^ this^study^demonstrated^ that^ lentiviral^vectors^pseudotyped^with^CoCVg^glycoprotein^derived^ from^Cocal^virus^could^be^ produced^at^titers^as^high^as^VSV-G-pseudotyped^vectors,^and^exhibited^both^broad^tropism^ and^high^stability.^Importantly,^pseudotyping^with^CoCVg^also^confers^resistance^to^viral^in- activation^by^human^serum^(Trobridge^et^al.,^2010).^^ In^yet^another^embodiment,^the^vesicular^envelope^glycoprotein^used^for^pseudotyping^the^ lentiviral^vector^is^PyriVg^having^an^amino^acid^sequence^having^at^least^80%,^preferably^at^ least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^ or^100%^sequence^identity^to^SEQ^ID^NO:^190.^Optionally,^it^may^be^encoded^by^a^nucleic^ 40^ ^ acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^ least^96%,^at^ least^97%,^at^least^98%,^at^least^99%^or^100%^sequence^ identity^to^SEQ^ID^ NO:^191.^The^glycoprotein^ is^ thus^capable^of^mediating^ entry^ into^a^host^ cell^ that^ can^be^ entered^by^PyriVg^having^SEQ^ID^NO:^190.^PyriVg^is^derived^from^the^Piry^virus,^which,^like^ VSVG,^MARAV^and^COCV,^belongs^to^ the^genus^vesiculovirus.^Similar^ to^CoCVg,^PyriVg^ was^shown^to^be^more^ resistant^to^complement-mediated^inactivation^by^mammalian^sera^ than^VSVg^(Tijani^et^al.,^2018).^However,^the^cellular^surface^receptor^facilitating^the^entry^of^ Piry^virus^into^a^host^cell^is^currently^unknown.^ The^ lentiviral^vector^may^furthermore^be^pseudotyped^with^at^ least^one^surface^protein^of^ influenza^A^virus^and / or^at^least^one^surface^protein^of^respiratory^syncytial^virus.^ The^at^least^one^surface^protein^of^influenza^A^virus^is^selected^from^the^group^comprising^ surface^proteins^NA,^HA^and^M2.^ Influenza^A^hemagglutinin^(HA)^is^a^homotrimeric^glycoprotein^and^serves^as^a^viral^attach- ment^ factor^ that^can^bind^ to^sialic^acid^containing^receptors^at^the^surface^of^a^ target^cell.^ Once^the^virus^has^bound^to^the^cell,^the^target^cell^engulfs^the^virus^via^receptor-mediated^ endocytosis.^Within^the^endosome,^HA^undergoes^a^series^of^conformational^changes^in^re- sponse^to^a^reduction^in^the^pH,^which^results^in^a^refolding^of^HA^to^expose^a^hydrophobic^ fusion^peptide^that^is^part^of^the^N-terminus^of^one^of^its^subunits.^The^exposed^fusion^peptide^ acts^like^a^molecular^grappling^hook^that^inserts^itself^in^the^endosomal^membrane.^In^conse- quence,^the^endosomal^membrane^and^the^viral^membrane^are^brought^into^close^contact^to^ facilitate^membrane^fusion^so^that^the^viral^RNA^can^enter^the^cell^cytoplasm.^The^HA^surface^ protein^used^for^pseudotyping^of^the^lentiviral^vector^may^have^an^amino^acid^sequence^hav- ing^at^least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^ 97%,^at^least^98%,^at^least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^192.^Optionally,^ HA^may^be^encoded^by^a^nucleic^acid^sequence^having^at^least^80%,^preferably^at^least^85%,^ at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^or^100%^ sequence^identity^to^SEQ^ID^NO:^193.^ Influenza^A^neuroaminidase^(NA)^is^an^enzyme^that^cleaves^sialic^acid^and^plays^an^important^ role^during^infection^of^a^host^cell^by^cleaving^sialic^acid^groups^to^which^HA^becomes^at- tached.^NA^furthermore^facilitates^the^release^of^newly^generated^viral^particles^from^the^in- fected^cell.^The^NA^surface^protein^used^for^pseudotyping^of^the^lentiviral^vector^may^have^ an^amino^acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^ 95%,^at^ least^96%,^at^least^97%,^at^ least^98%,^at^least^99%^or^100%^sequence^identity^to^ SEQ^ID^NO:^194.^Optionally,^NA^may^be^encoded^by^a^nucleic^acid^sequence^having^at^least^ 80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^ 98%,^at^least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^195.^ 41^ ^ Influenza^A^Matrix-2^(M2)^protein^is^a^proton^selective^viroporin^capable^of^assembling^into^a^ homotetrameric^proton^channel.^Once^the^virus^is^engulfed^by^its^target^host^cell,^M2^allowing^ passage^of^protons^into^the^virion^core^in^response^to^a^reduction^of^the^pH^within^the^host^ endosome.^The^acidification^of^the^interior^of^the^virus^facilitates^the^release^of^viral^ribonu- cleoparticles^into^the^host^cell’s^cytoplasm^upon^membrane^fusion.^The^M2^surface^protein^ used^for^pseudotyping^of^the^lentiviral^vector^may^have^an^amino^acid^sequence^having^at^ least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^ least^98%,^at^least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^196.^Optionally,^M2^may^ be^encoded^by^a^nucleic^acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^least^ 90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^or^100%^sequence^ identity^to^SEQ^ID^NO:^197.^ The^at^least^one^surface^protein^of^respiratory^syncytial^virus^(RSV)^is^selected^from^the^group^ comprising^surface^proteins^SH,^F^and^G.^ RSV^small^hydrophobic^(SH)^protein^functions^as^a^viroporin^and^can^alter^membrane^per- meabilty^in^the^host.^It^was^found^to^form^oligomers^that^behave^as^cation-selective^channels^ in^the^host^cell^membrane^(Gan^et^al.,^2012).^The^SH^surface^protein^used^for^pseudotyping^ of^the^lentiviral^vector^may^have^an^amino^acid^sequence^having^at^least^80%,^preferably^at^ least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^ or^100%^sequence^identity^to^SEQ^ID^NO:^198.^Optionally,^SH^may^be^encoded^by^a^nucleic^ acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^ least^96%,^at^ least^97%,^at^least^98%,^at^least^99%^or^100%^sequence^ identity^to^SEQ^ID^ NO:^199.^ The^RSV^Fusion^(F)^protein^is^embedded^in^the^viral^membrane,^and,^as^the^name^implies,^ facilitates^ fusion^with^ the^membrane^of^ a^ target^ host^ cell.^ The^F^ surface^protein^ used^ for^ pseudotyping^of^the^lentiviral^vector^may^have^an^amino^acid^sequence^having^at^least^80%,^ preferably^at^least^85%,^at^least^90%,^at^least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^ at^least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^200.^Optionally,^F^may^be^encoded^ by^a^nucleic^acid^sequence^having^at^ least^80%,^preferably^at^ least^85%,^at^ least^90%,^at^ least^95%,^at^least^96%,^at^least^97%,^at^least^98%,^at^least^99%^or^100%^sequence^identity^ to^SEQ^ID^NO:^201.^^ The^RSV^G^protein^ serves^as^an^attachment^glycoprotein^of^RSV.^G^ is^expressed^on^ the^ surface^of^ the^ virus^and^ facilitates^ binding^ of^RSV^ to^ the^CX3CR1^chemokine^ receptor^ in^ human^airway^epithelial^cells^(Johnson^et^al.,^2015).^The^G^surface^protein^used^for^pseudo- typing^of^the^lentiviral^vector^may^have^an^amino^acid^sequence^having^at^least^80%,^prefer- ably^at^least^85%,^at^least^90%,^at^least^95%,^at^ least^96%,^at^least^97%,^at^least^98%,^at^ least^99%^or^100%^sequence^identity^to^SEQ^ID^NO:^202.^Optionally,^G^may^be^encoded^by^ a^nucleic^acid^sequence^having^at^least^80%,^preferably^at^least^85%,^at^least^90%,^at^least^ 42^ ^ 95%,^at^ least^96%,^at^least^97%,^at^ least^98%,^at^least^99%^or^100%^sequence^ identity^to^ SEQ^ID^NO:^203.^ Accordingly,^in^one^aspect,^the^composition^for^use^according^to^the^present^invention^com- prises^a^lentiviral^vector^that^is^pseudotyped^with^VSVg^and^Influenza-NA.^ In^another^aspect,^ the^composition^for^use^according^ to^the^present^ invention^comprises^a^ lentiviral^vector^that^is^pseudotyped^with^VSVg^and^Influenza-HA.^In^another^aspect,^the^com- position^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudo- typed^with^VSVg^and^Influenza-M2.^In^another^aspect,^the^composition^for^use^according^to^ the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg^and^In- fluenza-NA.^In^another^aspect,^ the^composition^ for^use^according^ to^the^present^ invention^ comprises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg^and^Influenza-HA.^In^another^ aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vec- tor^that^is^pseudotyped^with^MaraVg^and^Influenza-M2.^In^another^aspect,^the^composition^ for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^ with^CoCVg^and^Influenza-NA.^In^another^aspect,^the^composition^for^use^according^to^the^ present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg^and^Influenza- HA.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^comprises^ a^lentiviral^vector^that^is^pseudotyped^with^CoCVg^and^Influenza-M2.^In^another^aspect,^the^ composition^ for^use^according^to^the^present^ invention^comprises^a^ lentiviral^vector^that^ is^ pseudotyped^with^PyriVg^and^Influenza-NA.^In^another^aspect,^the^composition^for^use^ac- cording^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^PyriVg^ and^Influenza-HA.^In^another^aspect,^the^composition^for^use^according^to^the^present^inven- tion^comprises^a^lentiviral^vector^that^is^pseudotyped^with^PyriVg^and^Influenza-M2.^In^another^ aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vec- tor^that^is^pseudotyped^with^VSVg^and^RSV-SH.^In^another^aspect,^the^composition^for^use^ according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^VSVg^ and^RSV-F.^In^another^aspect,^ the^composition^for^use^according^to^the^present^ invention^ comprises^a^lentiviral^vector^that^is^pseudotyped^with^VSVg^and^RSV-G.^In^another^aspect,^ the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^with^MaraVg^and^RSV-SH.^ In^another^aspect,^ the^composition^for^use^ac- cording^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg^ and^RSV-F.^In^another^aspect,^ the^composition^for^use^according^to^the^present^ invention^ comprises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg^and^RSV-G.^In^another^aspect,^ the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^with^CoCVg^and^RSV-SH.^In^another^aspect,^the^composition^for^use^accord- ing^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg^and^ RSV-F.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^com- prises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg^and^RSV-G.^In^another^aspect,^the^ composition^ for^use^according^to^the^present^ invention^comprises^a^ lentiviral^vector^that^ is^ 43^ ^ pseudotyped^with^PyriVg^and^RSV-SH.^In^another^aspect,^the^composition^for^use^according^ to^ the^present^ invention^comprises^a^ lentiviral^vector^ that^ is^pseudotyped^with^PyriVg^and^ RSV-F.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^com- prises^a^lentiviral^vector^that^is^pseudotyped^with^PyriVg^and^RSV-G.^ In^further^aspects,^the^lentiviral^vector^is^pseudotyped^with^a^vesiculoviral^envelope^glycopro- tein,^at^least^one^surface^protein^of^respiratory^syncytial^virus^and^at^least^one^surface^protein^ of^influenza^A^virus.^ For^ instance,^ the^ composition^ for^ use^according^ to^ the^present^ invention^may^comprise^a^ lentiviral^vector^that^is^pseudotyped^with^VSVg,^Influenza-NA^and^RSV-SH.^In^another^aspect,^ the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^with^VSVg,^Influenza-NA^and^RSV-F.^In^another^aspect,^the^composition^for^ use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^ VSVg,^Influenza-NA^and^RSV-G.^In^another^aspect,^the^composition^for^use^according^to^the^ present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^VSVg,^Influenza-HA^ and^RSV-SH.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^ comprises^a^ lentiviral^vector^ that^ is^pseudotyped^with^VSVg,^ Influenza-HA^and^RSV-F.^ In^ another^aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^len- tiviral^vector^that^is^pseudotyped^with^VSVg,^Influenza-HA^and^RSV-G.^In^another^aspect,^the^ composition^ for^use^according^to^ the^present^ invention^comprises^a^ lentiviral^vector^that^is^ pseudotyped^with^VSVg,^Influenza-M2^and^RSV-SH.^In^another^aspect,^the^composition^for^ use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^ VSVg,^Influenza-M2^and^RSV-F.^In^another^aspect,^the^composition^for^use^according^to^the^ present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^VSVg,^Influenza-M2^ and^RSV-G.^In^another^aspect,^ the^composition^for^use^according^ to^the^present^invention^ comprises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg,^Influenza-NA^and^RSV-SH.^In^ another^aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^len- tiviral^vector^that^is^pseudotyped^with^MaraVg,^Influenza-NA^and^RSV-F.^In^another^aspect,^ the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^with^MaraVg,^Influenza-NA^and^RSV-G.^In^another^aspect,^the^composition^ for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^ with^MaraVg,^Influenza-HA^and^RSV-SH.^In^another^aspect,^the^composition^for^use^accord- ing^to^ the^present^ invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg,^ Influenza-HA^and^RSV-F.^In^another^aspect,^the^composition^for^use^according^to^the^present^ invention^comprises^a^ lentiviral^vector^ that^ is^pseudotyped^with^MaraVg,^Influenza-HA^and^ RSV-G.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^com- prises^a^lentiviral^vector^that^is^pseudotyped^with^MaraVg,^Influenza-M2^and^RSV-SH.^In^an- other^aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^ vector^ that^ is^pseudotyped^with^MaraVg,^ Influenza-M2^and^RSV-F.^ In^another^aspect,^ the^ composition^ for^use^according^to^the^present^ invention^comprises^a^ lentiviral^vector^that^ is^ 44^ ^ pseudotyped^with^MaraVg,^Influenza-M2^and^RSV-G.^In^another^aspect,^the^composition^for^ use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^ CoCVg,^Influenza-NA^and^RSV-SH.^In^another^aspect,^the^composition^for^use^according^to^ the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg,^Influenza- NA^and^RSV-F.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^ comprises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg,^Influenza-NA^and^RSV-G.^In^ another^aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^len- tiviral^vector^that^is^pseudotyped^with^CoCVg,^Influenza-HA^and^RSV-SH.^In^another^aspect,^ the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^with^CoCVg,^Influenza-HA^and^RSV-F.^In^another^aspect,^the^composition^for^ use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^ CoCVg,^ Influenza-HA^and^RSV-G.^In^another^aspect,^the^composition^for^use^according^to^ the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg,^Influenza- M2^and^RSV-SH.^In^another^aspect,^the^composition^for^use^according^to^the^present^inven- tion^comprises^a^lentiviral^vector^that^is^pseudotyped^with^CoCVg,^Influenza-M2^and^RSV-F.^ In^another^aspect,^ the^composition^for^use^according^ to^the^present^ invention^comprises^a^ lentiviral^vector^that^is^pseudotyped^with^CoCVg,^Influenza-M2^and^RSV-G.^In^another^aspect,^ the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^with^PyriVg,^Influenza-NA^and^RSV-SH.^In^another^aspect,^the^composition^ for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^ is^pseudotyped^ with^PyriVg,^Influenza-NA^and^RSV-F.^In^another^aspect,^the^composition^for^use^according^ to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^PyriVg,^Influ- enza-NA^and^RSV-G.^In^another^aspect,^ the^composition^ for^use^according^ to^ the^present^ invention^comprises^ a^ lentiviral^ vector^ that^ is^pseudotyped^with^PyriVg,^ Influenza-HA^and^ RSV-SH.^In^another^aspect,^the^composition^for^use^according^to^the^present^invention^com- prises^a^lentiviral^vector^that^is^pseudotyped^with^PyriVg,^Influenza-HA^and^RSV-F.^In^another^ aspect,^the^composition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vec- tor^that^is^pseudotyped^with^PyriVg,^Influenza-HA^and^RSV-G.^In^another^aspect,^the^compo- sition^for^use^according^to^the^present^invention^comprises^a^lentiviral^vector^that^is^pseudo- typed^with^PyriVg,^ Influenza-M2^and^RSV-SH.^ In^another^ aspect,^ the^composition^ for^use^ according^ to^ the^ present^ invention^ comprises^ a^ lentiviral^ vector^ that^ is^ pseudotyped^ with^ PyriVg,^Influenza-M2^and^RSV-F.^In^another^aspect,^the^composition^for^use^according^to^the^ present^invention^comprises^a^lentiviral^vector^that^is^pseudotyped^with^PyriVg,^Influenza-M2^ and^RSV-G.^ The^ inventors^ surprisingly^ found^ the^ combinations^ VSVg / Influenza-NA / RSV-SH^ and^ MaraVg / Influenza-NA / RSV-SH^resulted^in^particular^high^transduction^efficiencies^in^lung^cut^ slices.^^ 45^ ^ Accordingly,^ in^a^ preferred^ embodiment,^ the^at^ least^ one^ surface^ protein^ that^ is^ used^ for^ pseudotyping^is^Influenza-NA.^In^consequence,^the^composition^for^use^of^the^present^inven- tion^preferably^comprises^a^lentiviral^vector^pseudotyped^with^a^vesiculoviral^envelope^glyco- protein^and^at^least^Influenza^NA.^ Alternatively,^the^at^least^one^surface^protein^that^is^used^for^pseudotyping^is^preferably^RSV^ SH.^In^consequence,^the^composition^for^use^of^the^present^invention^may^comprise^a^lenti- viral^vector^pseudotyped^with^a^vesiculoviral^envelope^glycoprotein^and^at^least^RSV-SH.^ In^a^more^preferred^embodiment,^the^composition^for^use^of^the^present^invention^may^com- prise^a^lentiviral^vector^pseudotyped^with^a^vesiculoviral^envelope^glycoprotein,^Influenza-NA^ and^RSV-SH.^ The^inventors^showed^particular^advantages^if^the^composition^for^use^of^the^present^inven- tion^comprises^a^lentiviral^vector^pseudotyped^with^VSVg,^Influenza-NA^and^RSV-SH.^ Alternatively,^the^composition^for^use^of^the^present^invention^preferably^comprises^a^lentiviral^ vector^pseudotyped^with^MaraV-G,^Influenza-NA^and^RSV-SH.^ In^further^embodiments,^the^composition^for^use^of^the^present^invention^comprises^a^lentiviral^ vector^that^is^pseudotyped^as^described^in^the^Table^2^below:^ Table^2:^Combinations^of^different^pseudotypes^for^construction^of^lentiviral^vectors^ V g V .- ;^ - - - - - .- .- .- .- - - .- NA / Infl.-M2 / RSV-SH / RSV-G;^ Infl.-HA / Infl.-NA / Infl.-M2 / RSV- 46^ ^ SH / RSV-F;^ Infl.-HA / Infl.-NA / Infl.-M2 / RSV-F / RSV-G;^ Infl.-HA / Infl.- M .- ;^ - - - - - .- .- .- .- - - .- - .- C .- ;^ - - - - - .- .- .- .- - - .- - .- NA / Infl.-M2 / RSV-SH / RSV-G / RSV-F^ 47^ ^ PyriVg^ Infl.-HA / Infl.-NA;^ Infl.-HA / Infl.-M2;^ Infl.-NA / Infl.-M2;^ Infl.-HA / Infl.- ;^ - - - - - .- .- .- .- - - .- - .- ^ The^lentiviral^vector^of^the^composition^for^use^according^to^the^invention^furthermore^com- prises^a^cargo^sequence,^wherein^expression^of^said^cargo^sequence^in^a^cell^of^a^lung^may^ cure,^ameliorate^or^stabilize^a^respiratory^disease.^The^cargo^sequence^is^operably^linked^to^ the^internal^promoter / enhancer^region^described^in^detail^above.^It^comprises^a^nucleic^acid^ selected^from^the^group^comprising^a^protein-coding^gene,^a^miRNA,^an^shRNA^or^a^lncRNA.^^ ^ In^a^particularly^preferred^embodiment,^the^lentiviral^vector^of^the^composition^for^use^of^the^ invention^comprises^a^nucleic^acid^or^ cargo^sequence^which^ encodes^one^or^more^of^ the^ herein^disclosed^RNA^molecules^capable^of^ inducing^RNAi-mediated^immunity^against^hu- man^parainfluenza^virus,^i.e.^it^encodes^one^or^more^of^the^herein^disclosed^RNA^molecules^ comprising^a^nucleic^acid^sequence^of^any^of^SEQ^ID^NO:^1^to^56,^preferably^any^of^SEQ^ID^ NO:^1,^2,^3,^9,^12,^14,^15,^17,^18,^22,^23^and^29^and^most^preferably^any^of^SEQ^ID^NO:^1^or^ 23.^ ^ Throughout^ the^ invention,^ the^ term^ “about”^ is^ intended^ to^ be^ understood^ as^ ”+ / -^ 10%”.^ If^ “about”^relates^to^a^range,^it^refers^to^both^lower^and^upper^limits^of^the^range.^“A”^is^intended^ to^mean^“one^or^more”,^if^not^explicitly^mentioned^otherwise.^ ^ 48^ ^ All^literature^cited^herein^is^herewith^fully^incorporated.^The^present^invention^is^further^illus- trated,^but^not^limited,^by^the^following^examples.^ ^ Brief^description^of^the^Drawings^ ^ Fig.^1^ Knockdown^efficiency^of^lentivirally^encoded^shmiR30a^candidates^(A,^B)^and^ of^ lentivirally^encoded^shmiR^multiplex^candidates^(C,^D,^E)^ in^an^mTagBFP^ reporter^assay.^ Fig.^2^ ^ Knockdown^efficiency^of^siRNA^candidates^^in^an^mTagBFP^reporter^assay.^ Fig.^3^ Efficiency^of^ lentivirally^encoded^shmiR30a^candidates^in^an^HPIV3^infection^ model^in^LLCMK2^cells.^ Fig.^4^ Efficiency^of^siRNA^candidates^in^an^HPIV3^infection^model^in^LLCMK2^cells^ pre^and^post^infection.^ Fig.^5^ Efficacy^ and^ safety^ of^ post-exposure^ siRNA^ treatment^ of^ LLCMK2^ cells^ for^ HPIV3^infection.^ Fig.^6^ Efficacy^and^safety^of^post-exposure^siRNA^treatment^of^Calu-3^cells^for^HPIV3^ infection.^ Fig.^7^ Efficiency^of^ lentivirally^encoded^shmiR30a^candidates^in^an^HPIV3^infection^ model^in^human^PCLS.^ Fig.^8^ Efficiency^of^transfected^siRNA^candidates^in^an^HPIV3^infection^model^in^hu- man^PCLS.^ Fig.^9^ Efficacy^and^safety^of^postexposure^ treatment^of^human^PLCS^for^HPIV3^ in- fection^with^siRNA^candidate^1.^ Fig.^10^ Reduction^of^the^viral^load^of^parainfluenza^by^5'^palmitylated^siRNA^ Fig.^11^ Efficiency^of^siRNA^candidates^in^an^HPIV3^infection^model^in^LLCMK2^cells^ using^lipofectamine,^a^LNP^formulation^or^directly^palmityl^(P)^-^or^cholersterol^ (C)^–^modified^siRNA^ Fig.^12^ In^vivo^efficacy^and^safety^of^palmityl-modified^siRNA^candidate^1^ in^HPIV3- infected^cotton^rats.^ Fig.^13^ Transduction^ efficiency^ of^ differentially^ pseudotyped^ lentiviral^ particles^ as- sessed^by^mCherry^expression^detected^by^fluorescent^microscopy.^ Fig.^14^ Schematic^setup^of^plasmids^used^for^vector^production^and^screening^assay^ of^differentially^pseudotyped^lentiviral^particles^in^Calu3^cells.^ Fig.^15^ Transduction^efficiency^of^VSVg / Influenza-NA / RSV-SH^and^MaraVg / Influenza- NA / RSV-SH^pseudotyped^lentiviral^particles^in^PCLS.^ ^ Example^1^ 49^ ^ ^ shmiR30a^candidate^generation^^ HPIV3^ genome^ (strain^ C^ 243)^ negative^ and^ positive^ RNA^ strands^ were^ fed^ into^ the^ splashRNA^tool^(http: / / splashrna.mskcc.org / )^and^30^candidates^for^the^positive^strand^and^ 26^for^the^negative^strand^were^generated^(listed^in^table^3^and^4).^For^incorporation^into^the^ shmiR30a^ backbone,^ primer^ pairs^ including^ the^ predicted^ antisense^guide^ sequence,^ the^ sense^passenger^ sequence,^ the^miR30^stem^loop^and^4^bp^ overhangs^ for^BsmBI^ cloning^ were^ designed^ and^ ordered^ (Table^ 5^ and^ 6).^ The^ miR30a-encoding^ lentiviral^ vector^ (pL40C.SFFV.mCherry.miR30N.Spacer.gb)^was^cut^open^with^BsmBI^restriction^enzyme^to^ release^the^spacer^harboring^the^BsmBI^recognition^sites^and^leaving^overhangs^complemen- tary^to^the^designed^primer^pairs.^Primer^pairs^were^annealed^and^ligated^into^the^shmiR30a^ backbone.^ Lentiviral^ vector^ production^ from^ the^generated^vectors^was^ performed^as^ de- scribed^elsewhere^(Daily,^2016,^doi:10.1186 / s12859-016-0930-z).^ ^ Table^3:^positive^(+)^strand^target^sequences^for^HPIV3^^ s 1 2 3 4 5 6 7 8 9 1 1 1 1 1 1 1 1 1 19^ FGene_1595_v2^ SEQ^ID^NO:^19^ 50^ ^ 20^ FGene_1625_v2^ SEQ^ID^NO:^20^ 2 2 2 2 2 2 2 2 2 3 ^ Table^4:^negative^(-)^strand^target^sequences^for^HPIV3^^ s n 1 2 3 4 5 6 7 8 9 1 1 1 1 1 1 1 _ _ tive_1568_v2^ 51^ ^ 17^ HPIV3_FGene_Negative_71_v2^ SEQ^ID^NO:^47^ 1 1 2 2 2 2 2 2 2 ^ Table^5:^primer^pairs^for^positive^(+)^strand^target^sequences^for^HPIV3^^ F L L L L L L L L L L L H H H H H FGene_1626_v2_FW^ SEQ^ID^NO:^89^ FGene_1626_v2_REV^ SEQ^ID^NO:^90^ 52^ ^ FGene_1618_v2_FW^ SEQ^ID^NO:^91^ FGene_1618_v2_REV^ SEQ^ID^NO:^92^ F F F P P P P P M M M M ^ Table^6:^Primer^pairs^for^negative^(-)^strand^target^sequences^for^HPIV3^^ F H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N _ _ _ _ _ _ HPIV3_LGene_^ SEQ^ID^NO:^139^ HPIV3_LGene_^ SEQ^ID^NO:^140^ 53^ ^ Negative_291_v2_FW^ Negative_291_v2_REV^ H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N H ^ N ^ siRNA^candidate^generation^^ To^compare^shmiR30a^to^small^interfering^RNAs^(siRNAs),^target^sequences^of^five^well^per- forming^shmiR30a^candidates^were^designed^as^analogous^siRNAs^(Table^7).^For^enhanced^ RISC^loading,^siRNA^guide^strand^was^27^bp,^passenger^strand^was^25^bp^long.^^ ^ Table^7:^positive^(+)^strand^targeting^siRNA^sequences^for^HPIV3^^ s 1 1 1 2 2 ( ene_ _v ) Q : Q : ^ Candidate^screening^in^an^mTagBFP^reporter^assay^ 54^ ^ For^initial^screening^of^RNAi^candidates^an^mTagBFB^reporter^assay^was^performed^as^de- scribed^earlier^(Adams^et^al.,^2017,^^10.1016 / j.biomaterials.2017.05.032).^Briefly,^22^bp^com- plementary^sequences^for^all^designed^candidates^were^cloned^en^bloc^ into^ the^UTR^of^a^ mTagBFP^mRNA^and^introduced^lentivirally^into^a^32D^cell^line.^Upon^successful^knockdown^ by^the^candidates,^the^complete^mRNA^would^be^degraded^which^would^be^visualized^by^a^ loss^of^mTagBFP^fluorescence.^ 32D^cells^carrying^ the^mTagBFP^ reporter^were^ transduced^with^ the^shmiR30a^candidates^ and^cultured^for^72^hours^to^enable^reporter^knockdown.^Afterwards,^mTagBFP^fluorescence^ was^assessed^by^flow^cytometry^and^knockdown^efficiency^was^calculated^(results^in^Fig.^1A^ and^B).^The^two^most^successful^candidates^of^each^gene^were^used^for^further^testing.^ ^ Polycistronic^line^up^of^different^miRNA^backbones^(shmiR^multiplexing)^in^different^combi- nations^may^be^performed^to^achieve^parallel^knockdown^of^several^ targets,^as^disclosed,^ e.g.,^ in^Liu^et^al.,^2022.^For^shmiR^multiplexing,^miR30a,^miR223^and^miR144^backbones^ were^cloned^into^in^a^single^gene^sequence^at^any^given^combination.^In^between^the^miRNA^ backbones,^spacer^regions^of^0-99^base^pairs^of^noncoding^RNA^may^be^introduced^into^the^ lineup^(Fig.^1C).^In^a^fluorescent^reporter^assay,^the^efficacy^of^the^indicated^lentivirally-en- coded^ multiplex^ shmiR^ constructs^ was^ tested^ against^ their^ respective^ single^ shmiR^ con- structs.^Knockdown^efficiency^for^two^targets^(Fig.^1D^and^E)^was^assessed^three^days^after^ transduction^with^the^shmiRs.^Multiplex^shmiRs^performed^equally^efficient^than^the^respec- tive^single^targeting^shmiRs.^ ^ For^assessment^of^siRNA^candidates,^293T^cells^carrying^the^mTagBFP^reporter^were^trans- fected^with^the^siRNA^candidates^using^Lipofectamine^RNAiMax^and^cultured^for^48^hours^to^ enable^reporter^knockdown.^Afterwards,^mTagBFP^fluorescence^was^assessed^by^flow^cy- tometry^and^knockdown^efficiency^was^calculated^(results^in^Fig.^2).^ ^ shmiR30a^candidate^screening^in^an^HPIV3^infection^model^in^LLCMK2^^ LLCMK2^cells^were^transduced^lentivirally^with^16^most^effective^shmiR30-cadidates^(Table^ 8)^three^days^prior^to^infection^with^a^multiplicity^of^infection^(MOI)^of^1^for^6^hours.^At^the^day^ of^infection,^cells^were^infected^with^HPIV3^(strain^C^243,^2*102^focus^forming^units^(FFU)^in^ 200^µL).^One^hour^post^infection,^the^HPIV3^supernatant^was^withdrawn^and^cells^were^cul- tured^for^40^hours^at^35°C.^Afterwards,^supernatants^were^collected^and^HPIV3^titers^deter- mined^by^using^a^focus^forming^assay^(FFA)^on^LLCMK2^cells^(results^in^Fig.^3).^ ^ Table^8:^target^sequences^of^candidates^used^for^testing^in^LLCMK2^cells^^ 55^ ^ (+)^strand^ (-)^strand^ N L L H H F F P P M M ^ siRNA^candidate^screening^in^an^HPIV3^infection^model^in^LLCMK2^^ LLCMK2^cells^were^ transfected^with^ the^siRNAs^using^ Lipofectamine^RNAi^Max^24^hours^ prior^to^or^24^hours^post^infection^with^concentrations^ranging^from^0.01^nM^to^0.1^µM.^At^the^ day^of^infection,^cells^were^infected^with^HPIV3^(strain^C^243,^2*102^focus^forming^units^(FFU)^ in^200^µL).^One^hour^post^ infection,^the^HPIV3^supernatant^was^withdrawn^and^cells^were^ cultured^for^40^hours^at^35°C.^Afterwards,^supernatants^were^collected^and^HPIV3^titers^de- termined^by^using^a^focus^forming^assay^(FFA)^on^LLCMK2^cells^(results^in^Fig.^4).^ ^ Efficacy^and^safety^of^post-exposure^siRNA^treatment^of^LLCMK2^cells^for^HPIV3^infection^ Fig.^5^A^and^B^show^dose-dependent^reduction^of^HPIV3^viral^load^by^siRNA^candidate^1^(si1^ comprising^a^sequence^of^SEQ^ID^NO:^169,^Fig.^5A)^^or^siRNA^candidate^23^(si23^comprising^ a^sequence^of^SEQ^ID^NO:^175,^Fig.^5B)^treatment.^LLCMK2^cells^were^infected^with^2x102^ focus-forming^units^(FFU)^HPIV3^and^treated^with^the^indicated^siRNA^candidates^24^h^post^ infection^via^lipofectamine^transfection.^24^h^after^treatment,^the^supernatant^of^the^cells^was^ collected,^and^the^viral^load^was^determined^via^focus-forming^assays^(FFA).^Non-linear^re- gression^and^IC50^calculation^were^performed^with^IC50(si1)^=^0.013^nM^(95%^Cl^=^[0.007^to^ 0.086^nM]),^R2^=^0.794^(Fig.^5A)^and^IC50(si23)^=^0.42^nM^(95%^CI^=^[0.040^to^3.920^nM]),^R2^ =^0.44^(Fig.^5B).^n^=^3.^Fig.^5^C^and^D^show^the^ toxicological^assessment^of^ treatment^of^ LLCMK2^cells^with^siRNA^candidates^1^and^23.^In^Fig.^5C,^cell^lysis^was^investigated^by^a^ lactate^dehydrogenase^(LDH)^release^assay.^To^do^this,^cell^culture^supernatant^was^ana- lyzed^24^h^post^treatment^for^LDH^release.^Results^were^normalized^to^a^fully^ lysed^Triton^ control.^Fig.^5D^shows^the^effects^of^the^indicated^siRNA^candidates^on^metabolic^activity^of^ the^tested^cells^by^assessing^metabolization^of^WST-1.^To^do^this,^the^metabolic^turnover^of^ 56^ ^ WST-1^was^analyzed^24^h^post^ treatment.^Results^were^normalized^ to^untreated^medium^ control.^n=3.^ ^ Efficacy^and^safety^of^post-exposure^siRNA^treatment^of^Calu-3^cells^for^HPIV3^infection^ Fig.^6A^shows^ the^determination^of^a^suitable^ infection^dose^ for^HPIV3^ infection^of^Calu3^ cells.^Calu-3^and^LLCMK2^cells^were^ infected^with^2x100,^2x101,^2x102,^2x103,^and^2x104^ FFU^HPIV3^C243^in^an^FFA^and^viral^foci^were^counted^36^h^post-infection.^Fig.^6B^shows^ dose-dependent^reduction^of^HPIV3^viral^load^by^siRNA^candidate^1^(si1)^^or^siRNA^candidate^ 23^(si23)^treatment.^Calu-3^cells^were^infected^with^2x104^FFU^HPIV3^and^treated^with^the^ indicated^siRNA^candidates^24^h^post^infection^via^lipofectamine^transfection.^24^h^after^treat- ment,^the^supernatant^of^the^cells^was^collected,^and^the^viral^load^was^determined^via^focus- forming^ assays^ (FFA).^ Non-linear^ regression^ and^ IC50^ calculation^ were^ performed^ with^ IC50(si1)^=^0.64^nM^(95%^Cl^=^[0.231^to^1.553^nM]),^R2^=^0.828^and^IC50(si23)^=^0.8^nM^(95%^ CI^=^[0.452^to^26.89^nM]),^R2^=^0.6579.^n^=^3.^Fig.^6^C^and^D^show^the^toxicological^assess- ment^of^treatment^of^Calu-3^cells^with^siRNA^candidates^1^and^23.^In^Fig.^6C,^cell^lysis^was^ investigated^by^a^lactate^dehydrogenase^(LDH)^release^assay.^To^do^this,^cell^culture^super- natant^was^analyzed^24^h^post^treatment^for^LDH^release.^Results^were^normalized^to^a^fully^ lysed^Triton^control.^Fig.^5D^shows^the^effects^of^the^indicated^siRNA^candidates^on^metabolic^ activity^of^the^tested^cells^by^assessing^metabolization^of^WST-1.^To^do^this,^the^metabolic^ turnover^of^WST-1^was^analyzed^24^h^post^treatment.^Results^were^normalized^to^untreated^ medium^control.^n=3.^ ^ Preparation^of^human^precision-cut^lung^slices^ Precision-cut^ lung^slices^ (PCLS)^were^prepared^as^ described^elsewhere^ (Danov^O^ et^al.,^ 2018^doi:^10.1371 / journal.pone.0207767).^Briefly,^human^explant^lung^tissue^was^filled^with^ agarose,^punched^and^cut^with^a^ tissue^slicer^ to^200^–^300^µm^thick^round^8mm^slices^in^ diameter.^^ ^ shmiR30a^ candidate^ screening^ in^ an^ HPIV3^ infection^model^ in^ human^ precision-cut^ lung^ slices^^ PCLS^cells^were^transduced^lentivirally^with^12^most^effective^shmiR30-cadidates^three^days^ prior^to^infection^with^a^multiplicity^of^infection^(MOI)^of^10^for^6^hours.^At^the^day^of^infection,^ PCLS^were^infected^with^HPIV3^(strain^C^243,^2*104^FFU^in^200^µL).^One^hour^post^infection,^ the^HPIV3^supernatant^was^withdrawn^and^PCLS^were^cultured^for^72^hours^at^35°C.^After- wards,^ supernatants^ were^ collected^ and^ HPIV3^ titers^ determined^ by^ using^ an^ FFA^ on^ LLCMK2^cells^(results^in^Fig.^7).^ ^ siRNA^candidate^screening^in^an^HPIV3^infection^model^in^human^precision-cut^lung^slices^^ PCLS^were^ transfected^with^ the^siRNAs^using^Lipofectamine^RNAi^Max^24^hours^prior^ to^ infection^with^concentrations^1^µM.^At^ the^day^of^ infection,^cells^were^infected^with^HPIV3^ 57^ ^ (strain^C^243,^2*104^FFU^in^200^µL).^One^hour^post^ infection,^the^HPIV3^supernatant^was^ withdrawn^and^PCLS^were^cultured^for^72^hours^at^35°C.^Afterwards,^supernatants^were^col- lected^and^HPIV3^titers^determined^by^using^a^focus^forming^assay^(FFA)^on^LLCMK2^cells^ (results^in^Fig.^8).^ ^ Efficacy^and^safety^ of^ post-exposure^si1^ treatment^of^human^ in^human^precision-cut^ lung^ slices^for^HPIV3^infection^ Fig.^9A^shows^dose-dependent^reduction^of^HPIV3^viral^load^in^h^PCLS^by^siRNA^candidate^ 1^(si1)^treatment.^hPCLS^were^infected^with^2x104^FFU^HPIV3^and^treated^24^h^post^infection^ via^lipofectamine^transfection.^48^h^after^treatment,^the^supernatant^of^the^cells^was^collected,^ and^the^viral^load^was^determined^via^FFA.^Non-linear^regression^and^IC50^calculation^were^ performed^with^IC50(si1)^=^127.2^nM^(95%^Cl^=^[53.60^to^315.4^nM]),^R2^=^0.8411.^n^=^5.^Fig.^ 9^B^and^C^show^the^toxicological^assessment^of^treatment^of^hPCLS^cells^with^siRNA^candi- date^1.^In^Fig.^9B,^cell^lysis^was^investigated^by^a^lactate^dehydrogenase^(LDH)^release^as- say.^To^do^this,^the^supernatant^was^analyzed^48^h^post^treatment^for^LDH^release.^Results^ were^normalized^to^a^fully^lysed^Triton^control.^Fig.^9C^shows^the^effects^of^siRNA^candidate^ 1^on^metabolic^activity^of^the^tested^hPCLS^cells^by^assessing^metabolization^of^WST-1.^To^ do^this,^the^metabolic^turnover^of^WST-1^was^analyzed^48^h^post^treatment.^Results^were^ normalized^to^untreated^medium^control.^n=3.^ ^ Example^2^ Covalent^lipid^conjugates^for^the^inhalation^application^of^antiviral^siRNA.^ ^ The^pulmonary,^topical^application^of^siRNA^by^inhalation^requires^the^use^of^suitable^formu- lations^or^RNA^modifications.^These^allow^the^siRNA^to^enter^the^cell^for^a^sufficient^pharma- cological^effect.^In^the^following,^the^inhalative^administration^of^antiviral^siRNAs^bearing^di- rectly^conjugated^palmityl^residues^at^different^positions^was^tested.^ ^ Reduction^of^parainfluenza^viral^load^by^5'^palmitylated^siRNA.^^ ^ For^the^results^shown^in^Fig.^10A,^LLCMK2^cells^were^infected^with^2*102^FFU^HPIV3.^24^h^ post-infection,^the^cells^were^treated^with^5'^palmitylated^siRNA^1(100^nM)^or^LNP-formulated^ siRNA^1^(siRNA^1^consists^of^a^guide^RNA^of^SEQ^ID^NO:^169^and^a^passenger^strand^of^ SEQ^ID^NO:^170)^and^the^viral^load^was^determined^24^h^after^treatment^(Fig.^10^A).^For^the^ results^shown^in^Fig.^10B,^LLCMK2^cells^were^infected^with^2*102^FFU^HPIV3^and,^24^h^post- infection,^treated^with^5'^palmitylated^siRNA1^(0.1-^1000^nM)^or^LipofectamineRNAiMax^for- mulated^siRNA1^(0.1-^1000^nM).^The^viral^load^was^determined^24^h^after^treatment.^For^the^ results^shown^in^Fig.^10C,^human^PCLS^were^infected^with^2*104^FFU^HPIV3^and,^24^h^post- 58^ ^ infection,^ treated^with^5'^palmitylated^siRNA^1^(1^pM)^or^LipofectaminRNAiMax^ formulated^ siRNA^1.^The^viral^load^was^determined^48^h^after^treatment.^ ^ It^was^found^that^100^nM^of^unformulated,^5'^palmitylated^siRNA1^was^able^to^reduce^the^viral^ load^in^the^same^way^as^nebulized^LNP-formulated^siRNA1^(Fig.^10A).^The^5'^palmitylated^ siRNA^was^further^able^to^reduce^the^parainfluenza^virus^load^in^a^dose-dependent^manner,^ as^was^the^in^vitro^transfection^reagent^lipofectamine^(used^as^in^vitro^positive^control,^Fig.^ 10B).^In^human^lung^slices,^the^unformulated^5'^palmitylated^siRNA^was^also^able^to^inhibit^ the^viral^load^in^the^same^way^as^the^positive^control^(Fig.^10C).^ ^ Efficiency^of^siRNA^candidates^in^an^HPIV3^infection^model^in^LLCMK2^cells^using^lipofec- tamine,^a^LNP^formulation^or^directly^palmityl^(P)^-^or^cholersterol^(C)^–^modified^siRNA^ 20.000^LLCMK2^cells^were^infected^with^2*102^FFU^HPIV3^and^treated^with^the^respectively^ modified^siRNAs^(comprising^a^sequence^of^SEQ^ID^NO:^169)^24^hours^post^infection^without^ further^formulation.^Viral^load^was^assessed^24^hours^post^treatment^and^compared^to^non-^ modified^siRNA^delivered^by^Lipofectamine^or^LNP^(Fig.^11).^N1,^N6^and^N23^ indicate^ the^ nucleotide^position^within^the^siRNA^to^which^the^palmityl^(P)^or^cholesterol^(C)^modification^ was^added.^ ^ Palmityl-conjugations^to^siRNA1^at^the^3'^end^or^at^the^sixth^nucleotide^from^the^5'^direction^ of^the^respective^passenger^strand^(Fig.^2,^N23^and^N6),^as^well^as^a^cholesterol^residue^at^ the^5'^position,^resulted^in^a^reduction^in^parainfluenza^virus^load^that^was^comparable^to^that^ observed^for^nebulized^LNP^formulations^and^the^lipofectamine^control.^ ^ In^vivo^efficacy^and^safety^of^palmityl-modified^siRNA^candidate^1^in^HPIV3-infected^cotton^ rats^ Fig.^ 12A^ shows^ the^ schematic^ design^ of^ the^ in^vivo^experiment^ conducted^ in^ cotton^ rats.^ Cotton^rats^were^infected^with^2x107^FFU^HPIV3^and^treated^on^day^zero^(d0),^day^1(d1)^and^ day^2^(d2)^with^30^µg / kg^(low^dose),^100^µg / kg^(medium^dose)^or^300^µg / kg^(high^dose)^of^ palmityl-modified^ siRNA^candidate^ 1^ having^ a^guide^strand^ comprising^a^ nucleic^ acid^ se- quence^of^SEQ^ID^NO:^169.^N1^indicates^the^nucleotide^position^within^the^passenger^strand^ of^the^siRNA^duplex^to^which^the^C16^palmityl^modification^was^added.^Animals^were^sacri- ficed^on^day^3^(d3)^post^infection.^Non-infected^and^infected^animals^treated^with^300^µg / kg^ palmityl-modified^scrambled^siRNA^were^used^as^controls.^siRNAs^were^administered^to^the^ animals^via^inhalation.^n^=^5.^Clinical^scores^(with^a^maximum^score^of^5)^and^relative^body- weight^of^infected^untreated^and^medium^dose^treated^animals^are^shown^in^Fig.^12^B^and^C.^ Fig.^12D^shows^viral^load^of^cotton^rat^lungs^three^days^post^infection^measured^by^qPCR.^ Fig.^12E^shows^the^percentage^of^lymphocyte^cells^from^cotton^rat^bronchoalveolar^lavage^ (BAL)^three^days^post^infection^assessed^by^differential^cell^count^staining.^ 59^ ^ ^ Example^3^ ^ Pseudotyping^of^lentiviral^particles^with^different^pseudotype^chymeras^ Initially,^ different^ lentiviral^ particles^ displaying^vesiculoviral^glycoprotein^pseudotypes^were^ screened^for^their^transduction^efficacy^on^PCLS^(Fig.^13)^On^this^basis,^pseudotypes^VSVg,^ MaraVg,^CoCVg^and^PyriVg^were^determined^to^serve^as^a^basis^for^further^pseudotype^chi- meras^to^be^generated.^ For^this,^RSV^surface^proteins^F,^G^and^SH^as^well^as^Influenza^A^surface^proteins^HA,^NA^ and^M2^were^incorporated^into^the^lentiviral^particle^surface^upon^vector^production^in^various^ combinations^(table^9).^Differentially^pseudotyped^vectors^were^screened^in^Calu3^lung^cells^ for^their^efficacy.^For^this,^transduction^was^performed^using^an^MOI^of^1^for^all^candidates.^ 48^hours^post^ transduction,^cells^were^analyzed^for^mCherry^expression^by^flow^cytometry^ (schematic^pseudotypes^and^results^in^Fig.^14).^The^most^successful^combinations^VSVg / In- fluenza-NA / RSV-SH^ and^MaraVg / Influenza-NA / RSV-SH^were^ further^ tested^ in^PCLS^with^ MOIs^of^1^and^10,^displaying^a^very^high^transduction^efficiency^and^expression^of^GFP^after^ transduction^in^these^human^lung^tissue^slices^(Fig.^15).^ ^ 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Shaw^and^Cornetta,^2014,^Design^and^Potential^of^Non-Integrating^Lentiviral^Vectors.^Bio- medicines^2(1),^14-35.^ Tijani^et^al.,^2018,^Lentivector^Producer^Cell^Lines^with^Stably^Expressed^Vesiculovirus^En- velopes.^Mol^Ther^Methods^Clin^Dev.^10,^303-312.^ Brown^KM,^Nair^JK,^Janas^MM,^Anglero-Rodriguez^YI,^Dang^LTH,^Peng^H,^Theile^CS,^Cas- tellanos-Rizaldos^E,^Brown^C,^Foster^D,^Kurz^J,^Allen^J,^Maganti^R,^Li^J,^Matsuda^S,^Stricos^ M,^Chickering^T,^Jung^M,^Wassarman^K,^Rollins^J,^Woods^L,^Kelin^A,^Guenther^DC,^Mobley^ 61^ ^ MW,^Petrulis^J,^McDougall^R,^Racie^T,^Bombardier^J,^Cha^D,^Agarwal^S,^Johnson^L,^Jiang^ Y,^Lentini^S,^Gilbert^J,^Nguyen^T,^Chigas^S,^LeBlanc^S,^Poreci^U,^Kasper^A,^Rogers^AB,^ Chong^S,^Davis^W,^Sutherland^JE,^Castoreno^A,^Milstein^S,^Schlegel^MK,^Zlatev^I,^Charisse^ K,^Keating^M,^Manoharan^M,^Fitzgerald^K,^Wu^JT,^Maier^MA,^Jadhav^V.^Expanding^RNAi^ therapeutics^to^extrahepatic^tissues^with^lipophilic^conjugates.^Nat^Biotechnol.^2022^ 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Claims

CLAIMS 1. An oligonucleotide molecule capable of targeting an RNA derived from human parain- fluenza virus, wherein the oligonucleotide molecule is a) an RNA molecule capable of inducing RNAi comprising a nucleic acid se- quence having at least 90% sequence identity to any of SEQ ID NO: 1 to 56, preferably to SEQ ID NO: 1, 18 or 23; or b) an antisense oligonucleotide comprising a nucleic acid sequence that corre- sponds at least to a subsequence of 10 or more nucleotides of any of SEQ ID NO: 1 to 56, preferably of SEQ ID NO: 1, 8 or 23.

2. The oligonucleotide molecule of claim 1, wherein the oligonucleotide is an RNA mol- ecule comprising a nucleic acid sequence of SEQ ID NO: 1 or an antisense oligonu- cleotide comprising a nucleic acid sequence of SEQ ID NO:

204.

3. The oligonucleotide molecule of claim 1, wherein the oligonucleotide is an RNA mol- ecule comprising a nucleic acid sequence of SEQ ID NO: 18 or an antisense oligonu- cleotide comprising a nucleic acid sequence of SEQ ID NO:

205.

4. The oligonucleotide molecule of claim 1, wherein the oligonucleotide is an RNA mol- ecule comprising a nucleic acid sequence of SEQ ID NO: 23 or an antisense oligonu- cleotide comprising a nucleic acid sequence of SEQ ID NO:

206.

5. A screening method for an oligonucleotide-based therapeutic for treating an infection with a respiratory virus of interest, the method comprising: a) identifying at least one target sequence in an RNA derived from the respiratory virus of interest; and b) generating one or more candidate oligonucleotide molecules comprising a nucleic acid sequence complementary to the at least one target sequence identified in step a), wherein the candidate oligonucleotide molecule is selected from the group consisting of an RNA molecule capable of inducing RNAi and an antisense oligo- nucleotide.

6. The method of any of claim 5, further comprising a screening step after step b) wherein the screening step comprises: i. cloning an RNA sequence identified in step a) or parts thereof into an UTR of an mRNA molecule encoding a fluorescent protein to obtain a reporter mRNA; ii. introducing said reporter mRNA into a cell,iii. introducing the one or more candidate oligonucleotide molecules generated in step b) into the cell, iv. Assessing the fluorescence intensity in the cells comprising the reporter mRNA to determine a knock-down efficiency for each candidate oligonucleotide mol- ecule.

7. The method of any of any of claims 5 or 6, further comprising a screening step after step b) wherein the screening step comprises: I. introducing the one or more oligonucleotide RNA molecules generated in step b) into a cell in cell culture, II. infecting the cell of (I) with the respiratory virus of interest, III. obtaining supernatant from the cell of (I) at a first time point and at least a second time point after infection; IV. Determining the virus titre in the supernatants obtained at the first time point and at the at least second time point of step (III), 8. The screening method of any of claims 5 to 7, wherein step a) is done in-silico and involves the use of an algorithm for identifying a suitable target sequence in the RNA and predicting candidate oligonucleotide molecules capable of complementary bind- ing to said target sequence, wherein the algorithm takes into consideration the ther- modynamic properties of the target sequence and the predicted candidate oligonucle- otide molecules.

9. The screening method of any of claims 6 to 8, wherein the method comprises an ad- ditional step after step b) and prior to the steps of claims 6 and / or 7, comprising aer- osolzing the candidate oligonucleotide molecule generated in step b), wherein the aer- osolized candidate oligonucleotide molecule is introduced into a cell of the screening step of any of claims 6 and / or 7.

10. A method of preparing an oligonucleotide-based therapeutic for treating an infection with a respiratory virus of interest, the method comprising steps of selecting a candi- date oligonucleotide molecule capable of targeting an RNA derived from a respiratory virus using the screening method of any of claims 5 to 9 and providing a vector com- prising or encoding said candidate oligonucleotide molecule to obtain the oligonucle- otide-based therapeutic for treating an infection with a respiratory virus of interest, wherein, optionally, the oligonucleotide-based therapeutic is formulated in a pharma- ceutical composition.

11. The screening method of any of claims 5 to 9 or the method of preparing an oligonu- cleotide-based therapeutic of claim 10, wherein the respiratory virus of interest is se- lected from the group comprising influenza virus, respiratory syncytial virus, parainflu- enza virus, metapneumovirus, rhinovirus, coronavirus, adenovirus, bocavirus, Ebola virus and Nipah virus.

12. An oligonucleotide-based therapeutic for treating an infection with a human parainflu- enza virus, wherein the therapeutic comprises one or more of the oligonucleotide mol- ecules of any of claims 1 to 4 or, if the oligonucleotide molecule is an RNA molecule capable of inducing RNAi, a nucleic acid encoding them, preferably a) a lentiviral vector comprising a nucleic acid encoding one or more of the RNA molecules of any of claims 1 to 4, b) a lipid nanoparticle comprising one or more of the oligonucleotide molecules of any of claims 1 to 4, c) a cationic polymer selected from the group comprising a PEI vector associated with one or more of the oligonucleotide molecules of any of claims 1 to 4, or d) one or more of the oligonucleotide molecules of any of claims 1 to 4 linked to a lipophilic moiety, preferably 2′-O-hexadecyl.

13. The oligonucleotide molecule of any of claims 1 to 4, the screening method of any of claims 5 to 9 or 11, the method of preparing an oligonucleotide-based therapeutic of any of claims 10 or 11 or the oligonucleotide-based therapeutic of claim 12, wherein the oligonucleotide molecule is an RNA molecule capable of inducing RNAi selected from the group comprising small interfering RNAs, short hairpin RNAs and artificial microRNAs.

14. The oligonucleotide molecule of claim 13, the screening method of claim 13, the method of preparing an oligonucleotide-based therapeutic of claim 13 or the oligonu- cleotide-based therapeutic of claim 13, wherein the oligonucleotide molecule is a small interfering RNA consisting of an antisense strand and a complementary sense strand, wherein the sense strand comprises a 2′-O-hexadecyl conjugated to its 3’ end, its 5’ end and / or to the nucleotide at nucleotide position 6.

15. The oligonucleotide molecule of any of claims 1 to 4, the screening method of any of claims 5 to 9 or 11, the method of preparing an oligonucleotide-based therapeutic of any of claims 10 or 11 or the oligonucleotide-based therapeutic of claim 12, wherein the oligonucleotide molecule is an antisense oligonucleotide selected from the group comprising a DNA antisense oligonucleotide and a gapmer.

16. A composition for use in treating a respiratory disease in a subject, wherein said com- position is administered to the airways of the subject, wherein the composition com- prises a lentiviral vector pseudotyped with a vesiculoviral envelope glycoprotein and at least one surface protein of influenza A virus and / or at least one surface protein of respiratory syncytial virus, preferably, with Influenza NA.

17. The composition fur use of claim 16, wherein the lentiviral vector is pseudotyped with a vesiculoviral envelope glycoprotein, at least one surface protein of respiratory syn- cytial virus and at least one surface protein of influenza A virus.

18. The composition for use of any of claims 16 or 17, wherein the vesiculoviral envelope glycoprotein is selected from the group comprising MaraVg, VSVg, CoCVg and PyriVg, preferably, MaraVg or VSVg.

19. The composition for use of any of claims 16 to 18, wherein the at least one surface protein of respiratory syncytial virus is selected from the group comprising surface proteins SH, F and G, preferably, SH.

20. The composition for use of any of claims 16 to 19, wherein the at least one surface protein of influenza A virus is selected from the group comprising surface proteins NA, HA and M2, preferably, NA.

21. The composition for use of any of claims 16 to 20, further comprising a nucleic acid encoding one or more of the RNA molecules of any of claims 1 to 4.

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