Acid-labile Linker

For these reasons, an alternative route and more acid labile linkers compared to p-carboxy trityl linker 24a initially used were sought, to avoid high concentrations of TFA for the final cleavage. The synthesis of the alkoxysubstitued linkers 24b (Meisenbach and Voelter 1997) and 24c, which can be synthesised directly on the solid support in five steps, offer the possibility of linkers with tailor-made stability. 

The successful assembly of organic compounds on a solid support represents only part of the challenge in SPOS. After completion of synthetic sequence, the compounds must be cleaved from linkers attached to polymer by a chemical or photochemical reaction, for example, treatment of a polymer-bound compound with acids, bases, nucleophiles, redox reagents, and even photons. Acid-labile linker and amine-cleavable Marshall linker are two major classes of hnkers used in combinatorial synthesis. 

Many acid-labile linkers are used to assemble combinatorial libraries. Compounds are cleaved in the final step by TFA/DCM solution with various concentrations for a certain period of time. Mild cleavage conditions may lead to incomplete cleavage of the desired compound from a solid support. On the other hand harsh conditions may cause compound degradation and side reactions. Harsh conditions will also cause the partial breakdown of resin and the leaching of unidentified impurities into the final products. Harsh cleavage conditions demand the stabihty of all compounds under such conditions. This may limit the scope of combinatorial synthesis... 

For resin-bound carbamates (18-21), indole linker was the most acid-labile linker for this class of compounds. Rink linker ranked second. Resin (20) was cleaved with 0.5% TFA in 20 min. Resin (21) was cleaved with 1% TFA in 2 min. Resins (18) and (19) required higher concentration of TFA. Resin (18) was cleaved with 1% TFA in 5 h. Alternatively, resin (19) was cleaved with 1% TFA in 12 h and with 5% TFA in only 16 min (not shown). 

For resin-bound ureas (22-25), Indole and Rink linkers, generally ranked 1 and 2 in cleavage kinetics, were still the most acid-labile linker for this class of compounds. Resins (24) and (25) were cleaved with 0.5% TFA in 2 and 23 min, respectively. Resins (22) and (23) were cleaved with 1% TFA in more than 10 h. The order of cleavage rates is similar to carbamate compounds. 

The Marshall Unker [23] has been widely used to synthesize compounds that can be cleaved by primary and secondary amines to afford the corresponding amides. Marshall linker was used in the synthesis of three or more diversity-site hbraries because it allowed the addition of one more diversity element at the cleavage step. While the original reported linker [23] involved the oxidation of the Unker before cleavage, the efficient release of the resin-bound compounds using nucleophiles from the unoxidized linker has been reported [16, 24]. Similarly to the acid-labile linkers, the kinetics of the cleavage reaction and time required for this reaction directly affect the synthesis efficiency, purity and yield of the final products. A cleavage study was carried out on seven resin-bound thiophenol esters (34—40) on Marshall Unker with 3 amines (41-43) (Scheme 12.11 and Tab. 12.4). 

Chemical ligation methods for peptide synthesis using thioester chemistry in solution have been previously documented (see Vol. E 22a, Section 4.1.5). Generalized procedures for solid-phase ligation have been developed that simplify the overall procedure. One method uses a safety-catch acid labile linker at the C-terminus and was used for the synthesis of a 71-amino acid chemokine, vMIP I (Section 5.3.2.1). Another procedure uses a selectively cleavable glycolate ester linkage (Section 5.3.2.2). 

Two approaches for solid-phase chemical ligation have been described. Canne et al. have developed an elegant system that utilizes an oxime forming ligation to attach the first peptide to the resin, a selectively cleavable ester link to remove the peptide from the resin as a C-terminal carboxylic acid, and the Acm group to protect the N-terminal cysteine residue)311 A complementary approach has been developed by Brik et al. that utilizes native chemical ligation to attach the first peptide to the solid support, a safety-catch acid labile linker to remove the final polypeptide from the support as a C-terminal amide and either Acm or Msc group for N-terminal cysteine protection)32 ... 

Using the Safety-Catch Acid Labile Linker... 

The two main resin linkers developed so far are shown in Scheme 18, i.e. tris(alk-oxy)benzylamide- 412 and 4-alkoxybenzylamide-type linkers)341 the former being TFA labile and thus fully compatible with Fmoc/tBu and the latter strongly acid labile and correspondingly compatible with Boc/Bzl chemistry. As shown in the case of the tris(alk-oxy)benzaldehyde handle such handles may be introduced into the C-terminal amino acid ester by reductive amination, and after suitable N -protection coupled to amino-functionalized resins (see Scheme 18). Alternatively, the tris(alkoxy)benzaldehyde-functionalized resin, BAL resin, (see Scheme 14) is used to link the C-terminal amino acid ester by reductive amination. To overcome the difficult acylation of the V -arylamino acid ester derivative on resin (best results with 10 equivalent symmetrical anhydrides), synthesis in solution of the C-terminal dipeptide building block containing the amide handle followed by its attachment to the resin has been proposed)341 ... 

To avoid formation of piperazine-2,5-diones the use of mild acid labile N -protecting groups such as Trt or Ddz for the penultimate amino acid residue have been proposed in the case of the tris(alkoxy)benzyl linker, and subsequent acylation of the dipeptide as TFA salt. 

Figure 3.1. Acidolysis of a typical, acid-labile linker.

Acid-labile linkers are the oldest and still the most commonly used linkers for carboxylic acids. Most are based on the acidolysis of benzylic C-O bonds. Benzyl esters cleavable under acidic conditions were the first type of linker to be investigated in detail. The reason for this was probably the initial choice of polystyrene as an insoluble support for solid-phase synthesis [13]. Polystyrene-derived benzyl esters were initially prepared by the treatment of partially chloromethylated polystyrene with salts of carboxylic acids (Figure 3.3). 

Table 3.1. Acid-labile benzyl alcohol linkers. See text for additional cleavage reagents.

Dialkoxy- and trialkoxybenzyl esters are even more acid-labile than the Wang linker, and can, for example, be cleaved with dilute TFA, acetic acid, or hexafluoroiso-propanol without simultaneous acidolysis of Boc groups. These linkers thus enable the solid-phase synthesis of protected peptide fragments or other acid-sensitive products [33]. 

The trialkoxy benzhydrol linker, developed by Rink in 1987 [46] ( Rink acid resin , Figure 3.4) is a further acid-labile linker for carboxylic acids. Esters of this linker can, like trityl esters, be cleaved with acids as weak as acetic acid or HOBt [47], and care must be taken to avoid loss of the product during synthetic operations. 

Resin-bound (4-acyloxy-2-buten-l-yl)silanes, which can be prepared from resin-bound allylsilanes and allyl esters by cross-metathesis, react with dilute TFA to yield free carboxylic acids (Figure 3.7 [75]). However, the scope of this strategy remains to be explored. Similarly, esters of polystyrene-bound (2-hydroxyethyl)silanes readily undergo acidolysis and have been used as acid-labile linkers (Figure 3.7 [76]). 

Figure 3.7. (4-Acyloxy-2-buten-l-yl)silanes and 2-acyloxyethylsilanes as acid-labile linkers for carboxylic acids.

One of the oldest linkers for amides is the (4-methylbenzhydryl)amine linker (MBHA Entry 1, Table 3.11). In contrast to the corresponding benzhydrol linker (which is cleavable by 5% TFA in DCM, 5 min [45]), acidolysis of the benzylic C-N bond of the MBHA linker requires treatment with hydrogen fluoride or a similar acid. As for A-benzylamides, the acid-lability of Al-(diarylmethyl)amides increases with the number of electron-donating substituents on the aryl groups. 

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