CA2049361A1 - Oligodeoxynucleotides with 5'-linked chemical groups, method of product thereof and use thereof - Google Patents

Abstract

Method of synthesis is provided for novel oligodeoxynucleotides 5' linked to covalently attached chemical groups via intermediate linked phosphoramidites. The synthesis of the covalently attached chemical group to a phosphoramidite moiety occurs outside the automatic synthesizer, followed by application of this complex as a terminator of the 5' end of a phosphate-modified oligodeoxynucleotide in the automatic synthesizer. The compounds are used to attenuate or destroy mammalian gene expression or viral activity.

Description

WO 90/l2802 2 ~ ~ 9 3 6 ~ PCI/US90/01501 NOVE~ OLIGODEOXYN~CLEOTIDES WITH 5 ' ~ OED C}~CAI.
GRO~JPS, ~tOD OF PRODUCTION T~ EOF AND U5E T~ OF

'The ~ c:t i~lveQtion relates to novel oligod~3oxynucleotide~ 5' l~nlc2d to covalently a~ached c~emic~ oup~ via lnter~edlate linked phosphor~dltss, and the~ r m~thods o~ productiorl and u~ 5Ore speci~ically, th~ ect invent~on involves ~ synt,h~is o~ nthragllinon~ ~or other) linlced ~oUp to a phospho~a~sidito ~oiety out~lde ~ autor~tlc synt~esizer, ~ollowed by applicatlon o~ thi~3 co~pound as a ter~$nator o~ the 5 ~ ~nd o~ a phosphate-modi~l~d oligodeoxynucleotide ln thQ autoDIatic synthe~izer. The co~pounds are u~ed to attenuate or destroy ~a~calian gene ~xpre~ion or v~r~l activity.

~A~GR~I~IL5~ VE~5ION
Rec~nt dev~lop~en~s in molecular gsnetic~, such ~s t~o automated synth~sis of ollgodaoxynucl~otides and t~e us~ of computer 2C progra~s to calculata aoc~ssi~le mRN~ region~ as likely ta~g~ts for comple~entary b~se binding, p~ovide ~h~ ~asis for n~w ~pproaches to th~ control . o~ gana Qxpr~s~ion and ~e potsntlal ~or 'gene tha~:ap~ ' .
One approach has ~e~ to prepare a linXed psodu~t in which a chemothsrapeutic aqent ~s ~o~alently attached to 3ynth~tic ol~gonucleotides ~th sequences d~rected toward ~o~ple~entary DNA or ~RNA sequenc~s. The standard methad o~ preparing these llnXed products is to ~ynthesi~e normal-phosph~te oligonucleotides in ~he auto~atic 3ynt~esizer, ~ollowed by puri~ication o~ the 2~02 2 ~ ~ ~ 3 b ~ PCT/~Sgo/olSol -oligonucleotides and covalent attachment o~ the chemo~herapeut~c agent and/or li~ker group outside the auto~atic ~ynthes~zer. ~elene ~t al constructed oligodeoxynucleotlde~ with desivatized 3'-linXed inter~al~ting groups ~acrldines~ u~ing th~s ~tandard ~athod. ~slene, C., ~ont~n~y - Garestisr, T., Sai~on, ~. Qt al., OllqodeoxynuclQotides eovalantly ltnked ~o int~calatin~ ag~nts: ~ new cla3~ o~ 5~n~
regulatory ~ubstances. Biochemie, 67:777-783, 1985.

Desired propert~s o~ ~h~se linkad products includ~ ~lectiY~ suppr~ssion o~ genQ~axpress~on, stability against nucleasQq, and a balanc~ between a~ueou~ ~olubil~ty and membrane tr~nsportability.
Sev~ral ex~ples o~ the covalent linking o~
~ctl~e 9rOUp5 to ol~godeoxynuc3.eotides have been accomplished. For ex~pla, ra~earchQrs have devQloped synthes~ o~ lron ED~'A ch~lat~ng groups to normal phosphate oliqodeoxynucleotides co~ple~entary to ~peci~lc singl~ ~tranded DN~s. Thes~ can ~lve 20 ri3Q to reactivD oxygen-contain~ng radical~ in the ~i_inity Or the DNA. ~h~ co~pounds ~unction ~y ~l~av~q~ o~ ~h~ ~ingl~ stran~ed DNAs. So~e proble~s o~ no~m~l-phosphate oligo~eoxy~lucl~otid~s w~th linked E~TA cA~lating groups ~nclude: l) not 25 ach~e~ng co~pl~tQ ~p~cificity, 2) nonspec$~iclty in t~t ~h~ F~(II) O~ tha ~DTA ~ay not be tcthered to a ~inglo nucl~otid~ ~ite, 3~ autod~g~adatlon ln that ~h~ hlghly ~lci~nt clean~g~ reagPnts the~s~lves ~ay dest~oy th~ir own ~ac~bones, and 4) OH radicals 30 ~y di~use in ~ny d~rectio~s.
Rese~rchers hav~ also developed synthetic ~hods ~or linking alkylating groups to both the 3'-~n~s and 5' ends o~ nor~al phosphate oligonucleot~des. Rnorra et al attaohed a~ ;

.. . .

,, ~;
, W0 90/12802 2 ~ 4 ~ 3 ~ 1 PCM~S90/01501 irltercalating phenazine derivative l:o the 3 ' end of`
an ollgodeoxynuc:leotide ~rough an a~in~ lin}cQr, and an ~lkylating- group to the 5'ezld. RnDrr~ et ~1, Rezlctiv~ ol~gonuolQot~dQ der$~ ative~ and sQquence 5 sp~cli~ic mo~i~ic~tion oi~ ~uclelc acids. Bioc~emie, 67: 7850789, 1985 .
~ Pho~phzlte ~odi~ied oligodeoxynuclQotid~, suGh a~ pho~phorothioat~ ol~godeoxynuel~o~id~3 in which o~e o~ the noE~riding oxygen aton~ in ~ach 10 nucleotid~ i~ repl~ced by ~ ~ulfur ~to~, havQ the ~d~ant~g~os~ proplartie~ o~ good aqu~ous ~olubility, - and au~o~Dat~d s~3thesis via phosphora~nidites; and-~ave been noted ~or thelr antiviral e~ect.
Pho~ph~te-~odi~ied oligodeoxynucleot~des with 5'-covalently attached c~emical gr.oup~ vla intermedlate linked phosphoramidites h~ve not previously bsen o~tal~d.

~L~
NQthOd 0~ ~yntha~is ls ~provid~d ~or novel ol~godeoxynucleotide~ 5' llnked to covalently ~ttachad che~ical groups Yia l:nt~r~ediate linked phosphora~dites. Th~ synthesis o~ th~ covalently att~ched chem~G~l group to a p~osphorlmidlt~ ~oi~ty occur~ out i~Q th~ ~utomatic syn~h~sizer, ~ollowed by ~pplic~t~on of thi~ co~pl~x a~ a t~r~i~ato~ o~
5' ~nd o~ ~ phosph~t~-~od~gi~d ol~goleo~ynucleotide ln the ~uto~t~ synthesizer.
5~ co~pound~ are used to ~ttenuate or destroy ~ al~an ge~e ~xp~esslon or viral activity. Actual or pote~ial b~n2~its o~ the present inYent~on lnclud~ anti-~ N ~ctlv~ty in vit~o, ~pQCi~i~ity by ~ase ~equence o~ the tarqeted cell's g~nes rather than ~y ~ protein, improv~ sel~ctivity over : ~ : . . : . .
~, '' '~, , 1',,'., - :. . : : ~:.. , ,: - . , ;. ,-. . , ~.,':

WO50/1~02 PCT/US90/01501 -~936~

~on~ent~onal drug therapy and decrea~ed resistance, ~nd st~illty against nucleases.

The p~es~nt lnvention ~s directed to novel 5 ollgod~oxynucleot~d~s with 5'-linked che~ical groups, vla inter~diate linked pho~phoramidite3 and t~ ~ynth~s~s ~d U~Q ~her~o~. Th~o novul co~pound~ are d~rived by syn~h~3izing a pho~pho~am~dlt~ ~olaty ~o a coval~ntly li~c~d 10 ~h~o~cal group outsid~ an auto~atic ~ynthe~izer, ~ ~~ ~ ~ollow~d by puri~ication, ~nd application oi~ this phosphor~idite-covalently li~Xed group co~plex as a t~rminator o~ the 5~-end o~ a nor~al or phosphate-~odigied oligod~oxynucleot~de in an auto~ttc ~5 ~yn~h~ er.
~hls synthetlc ~ethod c~n pro~ide a wide ~ang~ oi' ~ensible co~pounds oi' the general ~ormula R-~rCDN, in whlch ODN i~ olige~deoxynucleotide, ~ is a llnker arm, and R is ~he cov~lently ~ttaahed 20 ~roup. ~n advAntaqeou~ 4x~mpl~ o~ a co~pound ~yn~hes~zed by thi~ ~e~hod is an~hraquinone 5'-ll~Xed to a phosphorothio~te oli~odeoxynuGleotlde a hydr~xyethyl-piperazinyl linXer arm, as ~en ~ n Exampl~ 1. Ano~her ~dvantageou~ example o~ a 25 ~ompo~nd syn~he~2~d by ~h~ ~ethod i~ p~osph~te-~odi~ied oligodaox~nu~looti~o wikh 5'-llnked acr~na unsu~t~tuted ~xmept ~or ~he 9-a~ino ~osition ~or l~nXag~ via a ~alei~ide ~nte~mediate, as ~en in Ex~mpl~ 2.
m Q co~ponents Or thes~ compounds o~ novel oligodeoxynucl~otides with 5'-linked hemical groups c~n be pr~pared a~ ~ollows.
The linker ~rm is an intermediate li~ked phosp~oramidite ~oiety. ~ prererred linker arm is a ,,, "

,. . . .: , . .
~ . '' ~ ', ' - :

~090/12802 2 ~ 4 9 3 ~ ~ PCT/US9OtO1501 piperazinyl derivative such as l-(2-hydroxyethyl) plp~raz~ne, although any linker ~sm which is co~YQni~nt ~nd appropr~ate ~or tha che~ical . conditions e~ployed in the ~yntheslzer ~ay be used.
For ~x~ple, ao2idi~e rlngs li~Xed via the 9-a~ino g~oup to oligos ~ prepared by ~elene et al are 8U~Ct to alkaline (a~onis) ~ydrolysis int he ~utc~at~c synthes~z~r, al~hough ~acondary ~m~nes are more 8ta~1Q than tPrtiary a~ines. In this ~tuation, ~he pr~r~d lin~er ar~ ls a ~leimide lnter~ediat~ to 5' link 9-a~lno 6-chloro acridine o~to an oligodeoxynucleotide. Thi 8ynth~is 0 malei~ide linked acrldine oligodeoxynucleotide is ln EY~mpl2 2. It allows the synthesis 0~
lS ollgodeoxynucleotides wlth all bases unb~ocXed by ~onia in the ~utomatl~ 3ynthesizer. Oth~rwi~e, vlthout thQ U~Q 0~ ~mmonia ona could obtain hlgh yield~ o~ 6-thiophenol substituted acridine only with norMal and phosphorothi.~te oligo-dT'~, which i~
tho only unhlocked b~sR.
~ covalently attachled ch2mical group are pre~rably anthraquinonQ or acridine. O~her covalently attached groups ~nclud~ intercalat~ng group~, known drugs ~uch as ~driamycln, bleo~yein, ~nd hydrolytic groups such as ~m~dazole. Another ~xa~ple wauld ~ ~nthraquinone attaehed via a poly~ethyloDa linker.
eoYalently ~ttac~ed g~oup 5'~ ked 9-~ino 6-c~loro acridine wa~ ~ound to be ~ub~ect to ~ubstitution at the C pos~tion by thiophenol used in t~ automatic syntheQ~z~r o de-me~hylate ~he ~ho~phot~ester product. To over~ome this problem, ~n a~rid~e unsubstituted except ~or the 9 positlon for linka~e was us~dO ~lso, a malei~ide :, : : :,, , ,:, ::~,- -: : .: , ...
,., ; : : .::
. . ~, . . ~ :
. . ... . .. .

intermediate linked the acridine onto the oligonucleotide. See previous discussion and Example 2.
Oligodeoxynucleotides of 5-30 mer, preferably 12-28 mer, are the reasonable and necessary lengths for hybridization that may be used in the present invention. Longer oligodeoxynucleotides are more expensive. Also, the oligodeoxynucleotide may be a thiol-containing oligodeoxynucleotide.
The oligodeoxynucleotide may be of different base sequences. Specific oligodeoxynucleotide base sequences may be synthesi2ed to complement regions of a viral DNA, viral RNA, mammalian DNA, or mammalian mRNA. The oligodeoxynucleotide is regarded as the means of delivering the covalently attached chemical group to the biological site in that it targets specific DNA or mRNA sequences.
The oligodeoxynucleotide can be either a normal oligo or a phosphate-modified oligo such as phosphorothioate or a co-polymer combination of both. Phosphate-modified oligodeoXynucleotide are preferred. Covalently linked groups have not previously been attached to the 5' end of phosphate-modified oligonucleotide.
A series of oligonucleotides (of oligo base sequences dTL, Td" dT~, dT~, dT", dC." antilsense c-myc 15-mer, and antisense anti-rev HIV sequence) were constructed and 5'-linked to acridine or anthraquinone via a hydroxyethyl-piperazinyl de~ivative. When phosphorothioate oligodeoxynucleotides were substituted in place of normal oligodeoxynucleotides, the result was more effective inhibition of HIV expression in an in vitro ~-cell assay.
The following non-limiting examples illustrate the invention in more detail.

':

- ~ , : ' , W0~0/12802 2 Q 4 9 3 ~ ~ P~T/US90/OIS01 ~ .

A ~ixture o~ l-ch~o~oanthraquinone tl.g~ and 1-(2-hydroxyethyl) plp~razine (5g) ~s heated at 150-~or 30 ~lnutes. ~fter cooling to room temperature, vat~r ~g added. 5he ~ixture is ~ilterQd. 1-tl-(2-~ydroxye~hyl)pip~raz~nyl~anthraquinone, ~.p. 168-(a~ter recrystallizatio~ ~rom C~Cl3) is obtained.
~-t1-t2 hydroxyothyl) plp~raz~yl~
ant~raquinone (336 my, 1 ffl~ol) ls dls~olvecl in C~2C12 - t2 ~1) and N-e~hyl-diisopropyla~ina ~760 p, 4 ~ol) i~ added. Then, N-N-diisopropyl-~ethyl-phosphoram~dlc chloride (194 ~, 4 ~mol) is added to the so:Lution. Arter 30 ~inutes, pour the solutlon into Q~hylac~tata (5 ml, previ.ously wa~hed wit~ 5%
N~C03) and oxtract with 2 x 5 ~l o~ 5% Na~C0~ and 2 x 5 ml Or saturated NaCl. Th~ ethyl acetate phase ~as dried over Na2S0~ and evaporat~d to an oil.
diisopropyl phosphamid~t~ ~di~-isopropylam~no pho3phate) 0-~thyl ester 18 obtain2d. ~he T~C on ~ilica gel ~athylacetate:trie1:hylamine - 9~ howed co~plete r~ction. R~ o~ th~ start~ng ~aterial-003, the productsO . 7 . ~lPNMR ~pectroscopy pPak o~
148. Without ~urther pur~ication, ~h~ o~l was dis~olved in th~ ~ppropriAt~ ~olum~ o~ aceto~itrile ~d u~ed d~roctly in Dliyonucleotide pr~p~ratlon.
Ths y~ld w~s ~bout 65%. {
Syn~hesis o~ phosphorothioate oligo~eoxynucleotid~ w~th the anti-~e~se 3~ti rev ~IV ba~e sequenee d(5'-TCG TCG CTG TCT CCG CTT CTT
CCT GCC ~) v~s c~rrled out usi~g s~andard phosp~ora~ldite t~chn~quas. ~n ~n ~uto~atic ~ynthesizer (an ABI 380B3, ~t~ach t~e diisopropyl phosphamidite (diisopropyl~mino phosphate) 0-~ethyl . ~ :: :: . : . .

~ - . ., . ~ . . ::
:- .- . ;

WO90/12~02 2 ~ 4 ~ 3 6 1 PCTIUS90/01501 o~ter to the 5' end o~ the phosphorothiGate oligod~oxynucleotide by ~eans o~ the standard ~ucleot~de ~ycl~. ~here is ~hus obta~ned 5'-~n~braquinons oligodeoxynucleotid , whioh is then deblocked and.pur~ied by ~PL~o ~3~

S-Trityl-3-~erc~ptoprop~nol(l~ol (334 ~g, l ~ol) wore dissolved iA ~2thylene chloride t2 2l~.
N-~thyldiisopropyl~mine (760 ~, 4 m~ol) was added, nnd methoxydiisopropylchloriophosphite (180~, l.2 ~mol) was slowly added ~t 0~. Tha m$xtures wero le~t ~or a ~urther 15 ~inutes at 0~. Th8 solutions ~ar~ poured into ethylscetate (5 ~l) (contained wlth t2iethylamina 0.5 ~l) and washed with 2x5ml o~ S%
Nn~C~ and 2x5ml o~ saturated NaCl. The organic phases w~re dried over Na~O~ and evaporated to oil. ^`
~9P NMR ~pectrum showed a s~ngle peak, ~146 and T~C
on silica gel (petroleum ether 9, triethyla~ine l:
o~ start~ng material 0.3, Rf o~ product 0.45).
~lthout gurther puri~ication, ~he clear oil was dissolved in dry acetonitrile for lO0 ~ solutisn.
~ypi~al yield wa3 about 40$.
Oligodeoxynualeotid~ synthe~i~ wa~ carrled out u~n~ standard solid-phase phosphora~ite techniques where~pon an ollgodaoxynucleotide co~ta~ing an S~t~iphenyl~e~hyl group attached to the 5' phosphate group via a car~on cha$n was o~ta~ned. 5he S-triphenyl~ethyl gTOUp is removed ~ith silver according to Connolly, B., 13 ~ucleic Acids ~esearch 44~5 (l985). Thiol-containing oligodeox~nucleotides are obtained.

.. . . .
~ ~, , ,::~' . ; , . :
, .:

~O90/12802 2 ~ ~ ~ 3 ~ ~ PCT/USgo/01~01 The ~$o~-containln~ olig~deoxynucleotides (1 p~l scale ~yn~he31s) w~ di~olv~d in 1~ Na~CO3 (2~1), and N (9-~Eidiny~ ide ~as added. N-(9-acridinyl) ~lei~ide h~d ~%en Qynthesized S ~ccord$ng to N~ra, Y. and Tuzi~ura , ~., 42 Aqric .
~iol. Ch~. 793, 1978. ~hQ ~iYtur~ w~ kQpt at ~e ~or 1~ houss, and th~n ~xtract~d ~th 3x2 ~1 o~
~hyla~at~t~. Tho~ is thu~ obtainad 2~ $de linXad acridine oligodeoxynucleotides. ~h~ ~xture ~aqu~ou~ ph ~) wa~ conc~ntrated w~th ~lash~d N2 g~s T~ produ~t~ ~ra ~urifi~d by ~P~. Y~eld~ Or 35-~0% wara o~t~ln~d ~o~ l~ol seale ~ynthesi~.

~hile pre~errad ~bodi~nts o~ the lnvention havo b~Q~ d~scr~b~d herein, lt w~ll bo evident to lS tho~Q ~Xllled ~n ~hQ art ~ro~ a reading o~ the ~o~go~ng dlsolosur~ o~ np~cl~ic oligodooxynucl~otidQ ba~ guQnces synthe~ized with 5'-llnkQd nnthra~u~nono or other che~ieal group~, ~hat ollgod~oxynuclQotida~ o~ di~ror~nt b~s ~quence~ can bo u~ed. Vnr~ous ohange~ and ~odl~lc~tions, aspecially pertaining to the ~oval~n~ly attachad chemicnl group5, may ~ ~a~e ~lt~out d~pa~ting rrO~ th~ ~pirit o~ th~ pre~e~t ntion.

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.::. ;. . - . ~ .

Claims (24)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE PROPERTY
OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS :

1. A compound, comprising an oligodeoxynucleotide linked via a linker arm to a group covalently attached to said linker, wherein said linkage of the oligonucleotide is effected via an intermediate of the linker arm that includes a phosphoramidite moiety; and said covalently attached group confers biological activity upon said compound.

2. The compound of claim 1, wherein the oligodeoxynucleotide is phosphorothioate.

3. The compound of claim 1, wherein said linker arm is lined to the 5' end of the oligonucleotide.

4. The compound of claim 3, wherein the linker arm is a piperazinyl derivative.

5. The compound of claim 3, wherein the covalently attached group is an intercalating group.

6. The compound of claim 3, wherein the covalently attached group is anthraquinone.

7. The compound of claim 3, wherein the covalently attached group is selected from the group consisting of interacalating compounds, chemotherapeutic drugs and hydrolytic compounds.

8. The compound of claim 3, wherein the oligodeoxynucleotide is a thiol-containing oligodeoxynucleotide, the covalently attached group is acridine, which is only substituted at the 9-amino position by covalent attachment to the linker arm, and the linker arm is maleimide.

9. The compound of claim 3, further comprising a second linker arm and covalently attached group, wherein said second linker arm and covalently attached group are linked to the oligonucleotide at the 3' end of said oligonucleotide.

10. The compound of claim 3, further comprising at least one additional linker arm that has a covalently attached group, wherein said additional linker arm and covalently attached group is linked to the oligonucleotide at nucleotides within the oligonucleotide chain.

11. A method of destroying or attenuating the activity of a virus in a mammal infected with said virus, comprising administering an effective amount of a compound of claim 3 to said mammal virus, wherein the nucleotide sequence of the oligonucleotide of said compound is sufficiently complementary to a targeted sequence of nucleotides of the DNA or-RNA-of said virus to bind to said sequence and thereby deliver said covalently attached group to said sequence; and said amount is sufficient to destroy or attenuate the activity of said virus.

12. A method of destroying or attenuating expression of a mammalian gene, comprising administering an effective amount of the compound of claim 3 to a mammal whose genome includes said gene, wherein the nucleotide sequence of the oligonucleotide of said compound is sufficiently complementary to at least a region of said gene or mRNA encoded thereby to bind to said gene or mRNA and thereby deliver said covalently attached group to said gene or mRNA in an amount that is sufficient to destroy or attenuate expression of said gene.

13. A method of inhibiting the expression of a human immunodeficiency virus (HIV) in vitro in T-cells, comprising introducing into said cells, an effective amount of a compound of claim 3, wherein the nucleotide sequence of the oligonucleotide of said compound is sufficiently complementary to at least a region the RNA of the said HIV or the proviral DNA
copy of said HIV to bind to said RNA or DNA; said covalently attached group is anthraquinone or acridine; and said amount is effective for inhibiting HIV expression.

14. The method of claim 13, wherein said nucleotide sequence is an antisense oligomer against rev.

15. The compound of claim 3, wherein said oligodeoxynucleotide includes phosphate-modified deoxynucleotide analogs.

16. The compound of claim 3, wherein said oligodeoxynucleotide is a copolymer that contains oligodeoxynucleotides and oligodeoxynucleotides that include phosphate-modified deoxynucleotide analogs.

17. The compound of claim 3, wherein the oligodeoxynucleotide is phosphorothioate.

18. A method for producing the compound of claim 3, comprising:
covalently binding the covalently attached group to the intermediate of linker arm that includes the phosphoramidite moiety to produce a second intermediate that includes the covalently attached group, the linker arm and the phosphoramidite moiety:
introducing said second intermediate into an automatic synthesizer that includes the synthetic oligonucleotide bound to a support, effecting attachment of said intermediate to the 5' end of said oligonucleotide by reacting said phosphoramidite moiety with the oligonucleotide, whereby said compound is produced, wherein said linker arm is selected such that the covalently attached group remains covalently attached via said linker arm in said synthesizer.

19. A method for producing the compound of claim 3, comprising:
synthesizing the oligonucleotide in the automatic synthesizer;

removing said synthetic oligonucleotide from the synthesizer; and covalently attaching a linker arm to said oligonucleotide by reacting the phosphoramidite moiety on an intermediate of the linker arm that includes the phosphoramidite moiety to link said arm with the 5' end of said oligonucleotide.

20. The method of claim 19, wherein the covalently attached group is linked to said linker arm prior to effecting attachment the linker arm to the oligonucleotide.

21. The method of claim 19, wherein the covalently attached group is linked to said linker arm subsequent to effecting attachment of the linker arm to the oligonucleotide.

22. The method of claim 19, wherein said oligonucleotide is a thiol-containing oligonucleotide, the covalently attached group is acridine, which is only substituted at the 9-amino position by covalent attachment to the linker arm, and the linker arm is maleimide.

23. The compound of claim 7, selected from the group consisting of adriamycin, bleomycin and imidazole.

24. A method for producing the compound of claim 3, comprising:
preparing an intermediate that includes a linker arm and a phosphoramidite moiety;
introducing said intermediate into an automatic synthesizer that includes the synthetic oligonucleotide bound to a support;
effecting attachment of said intermediate to the 5' end of said oligonucleotide by reacting said phosphoramidite moiety with the oligonucleotide, whereby an oligonucleotide with a linker arm bound thereto is produced;

removing said oligonucleotide with bound linker arm bound from the synthesizer and covalently attaching the covalently attached group so the linker arm to produce said compound.