Solid-Phase Synthesis and Combinatorial Technologies

								1.2	LINKERS 13
									O	Rdec
										N
			O						R2	H
										or
	N		NH2		R1				O	Rdec
					N
H		R				SPS	TFA/DCM			N
			1		H
N							1/1, 30', rt	R2	N	H
									H	or
1.11	O
									HN	Rdec
										N
								H2N		H
				O
				CbzNH		Cl	Cl
			NH2	R R	1		R1
	O		NH	2			N	R2	SPS
		O					NHCbz
	1.12
X							O
R3	Rdec		TFA/H2O/CH3CHO/TFE			CbzNH	X		X = OCO, O, NH
								R3
	Rdec
N			1/4/4/15, 4 hrs, rt			Rdec	Rdec
	NHCbz
							TFA/H2O/AcOH
	OH	R1	B(OH)2		O	SPS	18/1/1, 1 hr, rt			B(OH)2
N				N	B	R1		or		Rdec
	OH				O		THF/H2O

1.13

9/1, 2 hrs, rt

Figure 1.9 Acid-labile SP linkers 1.11–1.13.

The carbazate linker 1.12 (69), obtained from hydroxy ArgogelTM resin activated with CDI and reacted with anhydrous hydrazine, was used to support ketone-based protease inhibitors and to release them after SPS using TFA/H2O/CH3CHO/TFE 1/4/4/15 in 4 h at rt.

The diethanolamine linker 1.13 (70), obtained from aminomethyl PS resin and ethylene oxide, has been used to support boronic acids which, after SP transformations, were released with THF/H2O/AcOH 18/1/1 in 1 h at rt or, when acid-labile boronic acids were involved, with THF/H2O 9/1 in 2 h at rt.

This brief, incomplete survey should have provided a flavor of the many functional groups that can be hooked onto and released from a solid support using acid-labile linkers. Acids, alcohols, phenols, amines, hydroxylamines, and halides have been successfully attached to a variety of resins through the judicious choice of the linker

14 SOLID-PHASE SYNTHESIS: BASIC PRINCIPLES

that can be cleaved after elaboration to release acids, amides, sulfonamides, hydroxamic acids, and aromatics. The acidic cleavage conditions can be modulated according to the sensitivity of the final compounds, and the cleavage reagents are easily removed from the sample either by evaporation or, when necessary, by simple extraction. This is a significant advantage, particularly when many final compounds are simultaneously released and must be obtained in good purity with minimal purification, as is the case in combinatorial technologies.

1.2.3 Baseor Nucleophile-Labile Linkers

The wide use of acid-labile linkers and protecting groups in peptide SPS has reduced efforts toward the development of base-labile linkers. The commonly used SP Fmoc peptide coupling protocols require Fmoc deprotection under basic conditions during the synthesis, thus ruling out base-labile linkers. However, base-labile linkers are popular in oligonucleotide SPS and will be described in Section 2.2. Other examples of baseor nucleophile-labile linkers are shown in Fig. 1.10.

The fluorene linker 1.14 (71), which is easily attached to an aminomethyl PS resin, has been used to support C-terminal Boc-protected amino acids during oligopeptide synthesis. The cleavage of this linker requires the use of 20% piperidine, or better 20% morpholine at rt over 2 h.

The commercially available oxime linker 1.15 (72) has been used in peptide synthesis and for the SPS of small organic molecules such as indoles. Cleavage is carried out with either hydrazine (0.5 M hydrazine in CHCl3–MeOH 2/1 for 10 min at rt) or with aliphatic amines or amino esters (DCM at rt for 12 h) to produce hydrazides and amides, respectively.

The acetyldimedone linker 1.16 (73) is another example of a linker attached to an aminomethyl PS resin and has been used to support amines and amino acids during SPS. The products are released containing free amino groups. The cleavage conditions are typically 2% hydrazine–DMF for 5 min at rt.

The silicon linker 1.17 (74), prepared by Grignard reaction of a 4-bromoarylsilane with commercial formyl PS resin, was used to support acids, alcohols, and amines either as such via its acyl imidazole derivative; cleavage with TBAF (tert-butylam- monium fluoride) in DMF for 3 h at 60 °C, or with CsF in DMF for 18–24 h at 90 °C validated the release of the three different functionalities.

The p-thiophenol linker 1.18 (75), prepared from aminomethyl PS resin and 3-(4-thiophenyl)-propionic acid, was used to support a chloropyridazine and to release after SPS decorated aminopyridazines by treatment with primary or secondary amines and anilines for 24–48 h at 90 °C.

1.2.4 Photolabile Linkers

Photolabile linkers use light to break the bond between the elaborated intermediate and the linker, thus releasing pure compound from the SPS into solution without interference from potentially troublesome side products. This advantage has encour-

O
N	OH
H	COOH	SPS
NH R

1.14

O

NO2

COOH

R SPS

N

1.15HO

O
O	SPS
N	H
H
	N
	R

1.16O

1.2 LINKERS

15

H

N

	O
	O	O	HOOC
		Rdec	Rdec
		20%, DMF,
		2 hrs, rt
		NH2	O
	O		R1
		Rdec R1
	N		N Rdec
		O	H

H	N2H4	2%
N		Rdec
		H2N
Rdec
	DMF, 5', rt

Si	COOH		O		TBAF, DMF		COOH
	R1				3 hrs, 60°C	R1
		O	R1
					or		OH
						R2
					CsF, DMF
	XH		O
							NH2
OH	R2			R	18-24 hrs, rt	R2
	CDI	O	X	2
1.17		X = O, NH
SH
1.18		S	N			R2
				N	NHR1R2 R	N	N
		SPS				1	N
N O	Cl			Nu	24-48 hrs,		Nu
					90°C
H
	N
Cl
	N

Figure 1.10 Baseor nucleophile-labile SP linkers 1.14–1.18.

16 SOLID-PHASE SYNTHESIS: BASIC PRINCIPLES

aged the development of photolabile linkers for SPS, and some examples are shown in Fig. 1.11.

The original o-nitrobenzyl bromide linker 1.19 (76) attached to aminomethyl PS resin has been used for both peptide and small organic molecule SPS. Cleavage by photolysis at 350 nm under anaerobic conditions gives carboxylic acids; the insertion of an α-methyl allowed an easier cleavage (77). The linker 1.19 has also been prepared

	O	COOH
	NO2 R
		SPS
	N
	H
	Br

1.19

	O
	NO2	as for
	N
	H	1.19
	NHX

1.20 X=H 1.21 X=Me 1.22 X=Et

	O
	N	O	NO2	1) RCHO
	H		NH2 2) SPS
	1.23	OMe
			Me
	O	OH
	N	( )	ActivO SUGAR SPS
	H	4

1.24O2N

	O
N	OH	RCHO	O H
H				R
	OH		O

	NO2
		hν, 350nm	HOOC
				Rdec
		MeOH, 24h, rt
	O	Rdec
		O
	NO2	hν, 350 nm
			XHNOC
	TFE/DCM, 24 hrs, rt			Rdec
	N	Rdec
	X
	O
	OMe
	R		R	S
		hν, 350 nm
	S			R1
			HN
	N	R1 5% aq. DMSO,
	Me O	3 hrs, rt		O
	SUGARdec
	O
		hν, 350 nm		SUGARdec
		THF, 23 hrs, HO
	O2N
		rt

hν, 350 nm

RCHO benzene, 23 hrs, rt

1.25NO2

O

		OH
	O		COOH
		R	SPS	hν, 320 nm
					HOOC
		O
				O	Rdec
N		OH		THF, 10-30', rt
H	1.26			O Rdec

Figure 1.11 Photolabile SP linkers 1.19–1.26.

1.2 LINKERS 17

in the NH2 (1.20), NHMe (1.21), or NHEt (1.22) forms, which allow the synthesis of primary or secondary amides or C-terminal peptide amides (78, 79).

Another aminomethyl PS resin supported o-nitrobenzyl photolabile linker 1.23 (80) has been employed for the synthesis of heterocycles such as thiazolidinones. Attachment to the resin is through an acid or aldehyde group and photolytic cleavage is performed in 5% dimethyl sulfoxide (DMSO)–aqueous buffer to facilitate the biological testing of the final compound. A photolabile hydroxyl linker 1.24 (81), again supported on aminomethyl PS resin, has been used to synthesize carbohydrate and peptide derivatives in SP. The easily prepared diol linker 1.25 (82) allowed the attachment of aldehydes as acetals and their photo induced release using standard conditions.

A different, pivaloylglycol-based photolabile linker 1.26 (83) was prepared with a complex, eight-step procedure from dihydroxyacetone dimer and aminomethyl Tentagel resin; the linker was used to support and to further elaborate carboxylic acids. The photolytic release at 320 nm of elaborated acids compared favorably with more assessed o-nitrobenzyl linkers.

1.2.5 SAFETY-CATCH LINKERS

Some SP linkers are totally stable during the synthetic sequence and only become labile after a process known as activation, which increases the lability of the linker toward well-defined cleavage conditions. These linkers, known as safety-catch (SC) linkers, are very popular and allow the support and release of many different functionalities. Some examples that rely on different methods of activation are collected in Fig. 1.12 (1.27–1.30) and 1.13 (1.31–1.34).

The Kenner sulfonamide-based SC linker 1.27 was supported on PS resin (84) allowing the attachment of carboxylic acids or amino acids to the sulfonamide function. After synthetic elaboration, treatment with diazomethane produces the N-methylacylsulfonamide, which can be cleaved with nucleophiles such as 0.5 N NH3–dioxane or hydrazine–MeOH, 0.5 N NaOH, releasing amides, hydrazides, or carboxylic acids, respectively. A modification using iodoacetonitrile produces the more labile N-cyanomethyl derivative, which can be cleaved completely with stoichiometric amounts of amines to release the corresponding amides into solution.

The phenol-sulfide SC linker 1.28 (85) attached to Merrifield resin has been used to support peptides, with the exclusion of S-containing aminoacids. Oxidation of the sulfur atom with hydrogen peroxide increases the reactivity of the linker towards amines and allows the facile cleavage of the ester bond to release the final compounds into solution as amides.

The acid-labile SC linker 1.29 (86) bonded to aminomethyl PS resin, has been used to support acids and C-terminal Boc protected aminoacids. Reductive acidolysis (typically SiCl4–thioanisole–anisole/TFA at rt over 3 h) reduces the sulfoxide in the activation step and releases the free acid into solution because of the enhanced reactivity of the sulfide towards acid.

18 SOLID-PHASE SYNTHESIS: BASIC PRINCIPLES

	COOH	1) SPS	O	O	O		O
SO NH	R			S			Nucleophile
2	2			N		Rdec	Nuc Rdec
		2) Activation

1.27		I		X
		X
	X = Me or CH2CN
	OH				O		Rdec
								O
	COOH		1) SPS				R1 NH2 R
	R			S		O
S		2) Activation					1	N	Rdec
				O O				H
1.28			H2O2
O	O	OH		COOH		Activation/
					SPS	cleavage
N	N			R			HOOC
									Rdec
H	H					SiCl4/PhSH/
1.29
			S			PhOH/TFA,
			O			3 hrs, rt
		O
	OtBu				N		O
		N				N
O	O	N					OtBu
			COOH
				SPS			Activation/
N	N		R		O		cleavage	HOOC
H	H	OH		O	Rdec				Rdec
1.30							TFA, then
							pH 7

Figure 1.12 Safety-catch (SC) SP linkers 1.27–1.30.

The imidazole hydroxyl SC linker 1.30 (87) linked to hydrophilic PS resins has been employed for loading acids and C-terminal Fmoc-protected amino acids. Activation and deprotection of functional groups is performed by TFA treatment, which produces the imidazole TFA salt. The imidazole ring intramolecularly attacks the ester bond upon neutralization at pH 7 with aqueous buffer and releases the compound as the free acid into solution.

The thioketal-containing hydroxyl linker 1.31 (88) was prepared in three steps from Merrifield resin and used to support carboxylic acids. The stable resin-bound intermediate is activated via desulfurization, with either Hg(ClO4)2 or HIO4, and the resulting linker is photolyzed in standard conditions to give the pure, released acid.

The sulfide-based linker 1.32 (89), obtained from commercial thio-PEG–PS resin and chloropyrimidine, is activated to nucleophilic substitution via oxidation with perbenzoic acid after multistep SP transformations; treatment with amines then releases pure 2-aminopyrimidines in solution. Other nucleophiles should be suitable for the modular release of this and other heterocyclic S-supported nuclei.

The acetal linker 1.33 (90), obtained from suitably protected aminophenol (three steps from 2-nitro-5-methoxytoluene) and hydroxy PS resin, was activated by acetal hydrolysis to give acylindole derivatives which could be cleaved and diversified to give

1.2 LINKERS 19

					COOH
					1) R
			S		2) SPS			O	hν, 350 nm		HOOC
O
		S			3) Activation		O	Rdec			Rdec
1.31									THF/MeOH 3/1,
		OH			Hg(ClO4)2				2 hrs, rt
					or HIO4			O
CF								CF3			CF3
3	COOEt										R1
								R1
N							N		H		N
		SPS		Activation					N	R2 N N
S N								R3	R2
							S
			mCPBA, DCM,					N
						O	O	DCM,		R3
1.32				16 hrs, rt
								24 hrs, rt
					COOH			O	Rdec
	NH2			1)
					R			N			or
		OMe		2) SPS
O		OMe					O				or
1.33				3) Activation
				PPTS, PhCH3,
				16 hrs, 50°C
		O
R1R2NH				R
	Rdec		N	1					OH
THF, 72 hrs, rt
			R2
or											OH
MeOH/THF/NaNH2
			COOMe
	Rdec								()3
1/19/cat., 30', rt											1.34
or									OBn
NaOH1N/MeOH/dioxane		Rdec	COOH							1) HOOC-AA1-NHP
1/1/3, 16 hrs, rt
										2) Oxidative
										debenzylation
										3) Peptide SPS
										4) Activation
										NaBH4, THF,
O								OH		MeOH, 30', rt
					TBAF/THF						PEPT
	+	PEPT		COOH							O
()					N2, 20 hrs, rt
								()3
3

OH	OH

Figure 1.13 Safety-catch (SC) SP linkers 1.31–1.34.

20 SOLID-PHASE SYNTHESIS: BASIC PRINCIPLES

amides (amine, THF, 2 h, rt), esters (MeOH/THF 1/19, catalytic sodium amide, 30 min, rt) or acids (aqueous NaOH, MeOH/dioxane, 16 h, rt).

The redox-sensitive linker 1.34 (91), obtained in several steps from Merrifield resin and a lactone precursor, was charged with a N-protected aminoacid, treated with NBS to debenzylate and oxidize the linker to quinone, and submitted to SPS. The quinone linker was reductively activated to dihydroquinone with NaBH4 in THF/MeOH for 30 min at rt, then cleaved by treatment with anhydrous TBAF in THF for 20 h at rt to provide the free acidic peptide via intramolecular cyclization of the linker moiety.

1.2.6 Traceless Linkers

A major drawback to all of the examples encountered so far is the necessity of having a functional group in the final compound through which it is attached to the resin during the elaboration phase. This is true both for compounds that are directly attached to the resin and for compounds attached via a linker. In the latter case, the linker often leaves a residue on the cleaved compound; for example, the Rink amine linker leaves a terminal carboxamide residue. The search for so-called traceless linkers (TLs), that is, linkers that do not leave a residual functional group deriving from the cleavage reaction, has recently become a major area of interest in SP linker chemistry. These linkers are usually substituted with a hydrogen atom during the cleavage, but some alternative quenchers have also been used (vide infra). Some examples are reported in Figs. 1.14 (1.35–1.38), 1.15 (1.39–1.43) and 1.16 (1.44–1.46); additional references can be found in a recent review on TLs (92).

The silicon-based TL 1.35 (Fig. 1.14) (93) on aminomethyl PS resin has been used in the SP preparation of 1,4-benzodiazepines by decoration of the phenyl ring. Cleavage is effected by protodesilylation using the somewhat harsh anhydrous HF, which releases the unsubstituted phenyl. A more labile TL 1.36 (94) obtained by substituting Ge for Si can be cleaved with TFA–Me2S–water in the ratio 85/10/5. Many other silyl-based TLs have been reported (52–56, 92); cleavage conditions where H is replaced by I (ICl) or Br (Br2–pyridine) have been validated.

The phosphorus-based TL 1.37 (95), which can be prepared from the commercially available triphenylphosphine resin, can be cleaved to give various chemical functionalities depending on the cleavage conditions used. For example, a methyl group can be generated under strongly basic conditions, an alkene can be formed under Wittig reaction conditions, or alternatively, indoles can be obtained via the modified Madelung synthesis.

The hydroxymethyl resin-supported regenerated Michael (REM) linker 1.38 (96), has been used to support secondary amines during elaboration followed by release as tertiary amines by classical Hoffman elimination [DIEA/(diisopropyl ethylamine)– DMF at rt over 18 h] of the resin-bound quaternary ammonium salt.

The quinodimethane linker 1.39 (97) was easily prepared from hydroxymethyl PS resin and anthranilic acid; this linker can be used to perform hetero-Diels–Alder reactions to form condensed six-member heterocycles. Cleavage with Lewis acid– nucleophile cocktails in DCM for 16–24 h at rt produces unsubstituted or alkylsubstituted heterocycles in good yields.

							1.2	LINKERS		21
	O	O	X	SnMe3
			( )				COCl
	O		3		1) R1			SPS
N
				NHP
H	1.35	X=Si 1.36	X=Ge		2)		COOH
	P=Protecting Group				R2
	R3					R3
X		N	HF (1.35)				N
( )								R2
3		R2 TFA, 24 hrs, 60°C (1.36)
	N						N
	R1	O					O
					R1
Me	NaOMe,
H					KOtBu
	MeOH
N
	R1								R1
O	reflux,		PPh2+Br-		reflux, 45'			N
	4 hrs	1.37		H				H
					R1
				N
		CHO
		R2		O
		NaOMe, MeOH,
		R2	reflux
		H	R1
		N
		O
	O	H	X	O
		N								R1
	R1	SPS	R3	O			DIEA, DMF
O		R2			X-					N
									R3
1.38					R	1	18 hrs, rt			R2
				R3	N +
					R2

Figure 1.14 Traceless SP linkers 1.35–1.38.

The chromium carbonyl linkers 1.40 (98) and 1.41 (99) were prepared from commercial triphenylphospine resin and respectively from pre-formed p-arene chromium carbenes and Fischer chromium amino carbenes. Their SP elaboration is followed by cleavage with pyridine at reflux for 2 h (1.40) and with iodine in DCM for 1 h at rt (1.41); both linkers produce the desired compounds in good yields. A similar cobalt carbonyl linker 1.42 (100) was prepared as a mixture of mono- (1.42a) and bis- (1.42b) phosphine complex, either from pre-formed alkyne complexes on triphenylphosphine resin or by direct alkyne loading on the bisphosphine cobalt complex; traceless cleavage was obtained after SP transformations by aerial oxidation (DCM, O2, hn, 72 h, rt) and modified alkynes were released with good yields and

22 SOLID-PHASE SYNTHESIS: BASIC PRINCIPLES

		H	R1				R1
	R1	C				R2M, Nu
		X
O	105°-110°C,			X		DCM, rt,	X
1.39						16-24 hrs
		14 hrs			O		R2
							X=O, NTs
							R2=H, Me, Allyl,
							CH2COOtBu
	O						OH
PPh2Cr(CO)2		LiAlH4			pyridine
				OH
	1.40				2 hrs, reflux
MeO						MeO
PPh Cr(CO)				PPh2Cr(CO)4			O
2	4	NH2				I , DCM
		R				2	R
					R
						1 hr, rt	N
	OMe				N		H
1.41					H
				OH			OH
H					H
(OC)2 Co Co(CO)3				+	(OC)2 Co Co(CO)2
Ph2 P					Ph2 P	PPh2
1.42a						1.42b
				SPS

DCM, O2, hν, 72 hrs, rt

H Odec

Figure 1.15 Traceless SP linkers 1.39–1.42.

purities. A significant use of chromium and cobalt carbonyl complexes as TLs is to be expected for many solid-phase chemistry applications.

The carboxyl-based TL 1.43 (101) was easily prepared from hydroxymethyl PS resin and a trisubstituted aromatic compound; its SP functionalization on the amide carbonyl or on the chlorine atom is followed by cleavage with TMSI (trimethyl silyl iodide) for 72 h at 75 °C to obtain simultaneously ester hydrolysis and decarboxylation to 2-unsubstituted quinazolines. An expansion to other heterocyclic systems is easily foreseeable.