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Linkers for ADCs

Linkers for Bioconjugation: Linkers are an important part of a successful antibody-drug conjugate (ADC). Most ADCs are referred to with the payload and linker as one unit such as MC-MMAF, MC-Val-Cit-Pabc-MMAE, SPDB-DM4, and SMCC-DM1. Linker-payloads reflect how each evolved with the knowledge at the time. Linker technologies were associated with a payload class and were usually combined and used as such. However, both constructs are independent and there is no reason why, for instance, an auristatin cannot be coupled with a pH-sensitive linker or a disulfide linker. This section will describe options for linkers and how NJ Bio can help with selection of a linker for a selected payload.

Types of Linkers: Linkers are divided into cleavable and non-cleavable types. Cleavable linkers have a biological weak point that releases a metabolite either by reduction, hydrolysis, or proteolysis. Common cleavable linkers are MC-Val-Cit-Pabc, disulfides (e.g., SPDB, SPP), and hydrazone for pH-sensitivity. Many cleavable linkers use a self-immolating mechanism to release the intact payload. It is important to note that the pKa of the leaving group will be important to drive the self-immolation.

Disulfide linkers are also cleavable in the sense that they break in the presence of high levels of free cysteine or glutathione found in the cytosol of cells. Disulfides are interesting constructs, as the kinetics of release can be modulated by the steric hindrance around the disulfides. Another advantage of disulfides is that they can be attached directly to the cysteine of the antibody, resulting in a cleavable linker with fewer atoms added to the drug-linker complex. An example of this is SPDB-DM4. Newer disulfides have also been made to release alcohols.

Finally, examples of pH-sensitive linkers are usually found in hydrazone-based linkers that can hydrolyze in the more acidic environment of the lysosome. Carbonates have also been found to be pH-sensitive, as seen in the CL2A linker of SN38. The same release kinetics and metabolite in cells were shown with and without the cleavable sequence. The disadvantage of carbonates and hydrazones is that they do deconjugate in circulation. These linkers have received less attention because deconjugation can lead to toxicity. However, three currently approved ADCs have pH-sensitive linkers.

Cl2A Linker

Cl2A Linker

Hydrazone

Hydrazone

Non-cleavable linkers do not have a weak point and release from the antibody after complete lysosomal degradation of the ADC. Non-cleavable linkers are known for their stability and only release when the ADC completely degrades. The linker is part of the active metabolite as it stays attached to the payload and thus can change the property of the payload. A payload that has bystander activity can lose its bystander activity with a non-cleavable linker, which can also change the affinity of the payload to PGP pumps. Overall, non-cleavable linkers can change the activity of the active metabolite, which can impact activity, toxicity, and trafficking.

Finally, all types of linkers have had modifications to modulate the hydrophobicity, DAR and conjugation technology to create novel ADC constructs. Popular modified linkers are ones incorporating sugars, PEGS and enzymatic/click type conjugation. This additional solubility and DAR control can improve the therapeutic index by resulting in a better PK profile.

NJ Bio can design and select the right linker for your ADCs. Linkers can be mixed and matched with payload technology and optimized for your particular target, which can be a powerful way to enhance the overall success of your program. Contact NJ Bio to see how we can help with your linker needs.

 

The table below shows combinations of drug-linkers that have been used. Most of these can be adapted to different drug combinations, but these examples provide a visual representation of the structures of the most advanced and interesting drug-linkers.

mc-vc-PABC-MMAE ADC (Vedotin)

mc-vc-PABC-MMAE ADC (vedotin)

 

Clin. Cancer Res. 2012, 18, 5845–5849

https://DOI: 10.1158/1078-0432.CCR-12-1803

Blood 2018, 132 (Suppl. 1), 1683 DOI: 10.1182/blood-2018-99-118551

Cancer Res. 2016, 76, 3003–3013 DOI: 10.1158/0008-5472.can-15-1313

J. Clin. Oncol. 2018, 36, 3298–3306 DOI: 10.1200/JCO.2018.78.7697

Cancer Res. 2014, 74, 1214–1226 DOI: 10.1158/0008-5472.CAN-13-2440

Eur. J. Pharm. Sci. 2016, 93, 274–286 DOI: 10.1016/j.ejps.2016.08.015

Mol. Cancer Ther. 2014, 13, 2991–3000 DOI: 10.1158/1535-7163.MCT-13-0896

Nat. Med. 2018, 24, 203–212 DOI: 10.1038/nm.4472

Cancer Res. 2018, 78, 4059–4072 DOI: 10.1158/0008-5472.CAN-18-0327

SMCC-DM1 ADC (Emtansine)

SMCC-DM1 ADC (emtansine)

 

J. Med. Chem. 2014, 57, 6949–6964.  DOI: 10.1021/jm500766w

Future Oncol. 2013, 9, 319–326  DOI: 10.2217/fon.13.7

Adv. Med. Oncol. 2014, 6, 202–209 DOI: 10.1177/1758834014539183

N. Engl. J. Med. 2019, 380, 617–628 DOI: 10.1056/NEJMoa1814017

J. Exp. Clin. Cancer Res. 2016, 35, 104 DOI: 10.1186/s13046-016-0380-5

BMC Cancer 2019, 19, 517 DOI: 10.1186/s12885-019-5687-0

Invest. New Drugs 2018, 36, 869– 876 DOI: 10.1007/s10637-018-0570-4

mAbs 2019, 11, 1149–1161 DOI: 10.1080/19420862.2019.1618674

N-Acetyl-g-calicheamicin 1,2-dimethylhydrazine ADC (ozogamycin)

N-Acetyl-g-calicheamicin 1,2-dimethylhydrazine ADC (ozogamycin)

 

Clin Cancer Res. 2001, 7, 1490–1496

Clin Cancer Res 2018, 24, 3242–3246 DOI: 10.1158/1078-0432.CCR-17-3179

Oncologist 2018, 23, 1103–1108 DOI: 10.1634/theoncologist.2017-0604

Blood 2010, 116, 2618–2619 DOI: 10.1182/blood-2010-08-300871

Blood 2017, 130, 2373–2376 DOI: 10.1182/blood-2017-09-797712

Drugs 2017, 77, 1603–1610 DOI: 10.1007/s40265-017-0802-5

mc-MMAF ADC (mafodotin)

mc-MMAF ADC (mafodotin)

 

Clin. Cancer Res. 2018, 24, 4399–4406 DOI: 10.1158/1078-0432.CCR-18-0481

Blood Cancer J. 2019, 9, 37 DOI: 10.1038/s41408-019-0196-6

Blood 2014, 123, 3128–3138 DOI: 10.1182/blood-2013-10-535088

sulfo-SPDB-DM4 ADC (soravtansine)

sulfo-SPDB-DM4 ADC (soravtansine)

 

Mol. Cancer Ther. 2015, 14, 1605–1613 DOI: 10.1158/1535-7163.MCT-14-1095

SG3249 ADC (tesirine)

SG3249 ADC (tesirine)

 

Blood 2018, 131, 1094–1105 DOI: 10.1182/blood-2017-10-813493

Mol. Cancer Ther. 2016, 15, 2709–2721 DOI: 10.1158/1535-7163.MCT-16-0233

Cancer Res. 2018, 78 (13 Suppl.), 2792A DOI: 10.1158/1538-7445.AM2018-2792A

mc-GGFG-DXd ADC (durextecan)

mc-GGFG-DXd ADC (durextecan)

 

Clin. Cancer Res. 2016, 22,5097–5108 DOI: 10.1158/1078-0432.CCR-15-2822

Oncogene 2019, 38, 1398–1409  DOI: 10.1038/s41388-018-0517-4

J. Clin. Oncol. 2018, 36 (15 Suppl.), e24206 DOI: 10.1200/JCO.2018.36.15_suppl.e24206

CL2A-SN38 ADC (Govitecan)

CL2A-SN38 ADC (govitecan) 

 

Bioconjug. Chem. 2015, 26, 919–931 DOI: 10.1021/acs.bioconjchem.5b00223

Engl. J. Med. 2019, 380, 741–751 DOI: 10.1056/NEJMoa1814213

Clin. Genitourin. Cancer 2016, 14, e75–e79 DOI: 10.1016/j.clgc.2015.10.002

Clin. Cancer Res. 2015, 21, 3870–3878 DOI: 10.1158/1078-0432.CCR-14-3321

Mol. Cancer Ther. 2018, 17, 196–203 DOI: 10.1158/1535-7163.MCT-17-0442

Mol. Pharm. 2015, 12, 1836–1847 DOI: 10.1021/mp5006195

J. Clin. Oncol. 2017, 35, 2141–2148 DOI: 10.1200/JCO.2016.70.8297

vc-PABC-EDA-seco-DUBA ADC (duocarmazine)

vc-PABC-EDA-seco-DUBA ADC (duocarmazine)

 

Mol. Pharm. 2015, 12, 1813–1835 DOI: 10.1021/mp500781a

Mol. Cancer Ther. 2015, 14, 692–703  DOI: 10.1158/1535-7163.MCT-14-0881-T

SPDB-DM4 ADC (ravtansine)

SPDB-DM4 ADC (ravtansine)

 

Mol. Cancer Ther. 2015, 14 (12 Suppl. 2), C165.     DOI: 10.1158/1535-7163.MCT-18-0529

Mol. Cancer Ther. 2014, 13, 1537–1548. DOI: 10.1158/1535-7163.MCT-13-0926

Clin. Cancer Res. 2009, 15, 4028–4037. DOI: 10.1158/1078-0432.CCR-08-2867

J. Clin. Pharmacol. 2017, 57, 865–875. DOI: 10.1002/jcph.869

Cancer Res. 2017, 77 (13 Suppl.), 2630 DOI: 10.1158/1538-7445.AM2017-2630

β-Glucuronide MMAE ADC

β-Glucuronide MMAE ADC

 

Blood 2016, 128, 4470 DOI: 10.1182/blood.V128.22.4470.4470

AS269 ADC

AS269 ADC

 

Cancer Res. 2015, 75 (15 Suppl.), 639 DOI: 10.1158/1538-7445.AM2015-639

mc-vc-PABC-Aur0101 ADC

mc-vc-PABC-Aur0101 ADC

 

Sci. Transl. Med. 2017, 9, eaag2611 DOI: 10.1126/scitranslmed.aag2611

SGD1910 ADC (talirine)

SGD1910 ADC (talirine)

 

Bioconjugate Chem. 2013, 24 (7), 1256–63. DOI:10.1021/bc400217g

Sulfo-SPDB-DGN462 ADC

Sulfo-SPDB-DGN462 ADC

 

Mol. Cancer Ther. 2018, 17, 1271–1279  DOI: 10.1158/1535-7163.MCT-17-1077

DGN549 ADC

DGN549 ADC

 

Blood Adv. 2018, 2, 848–858 DOI: 10.1182/bloodadvances.2018017517

Mol. Cancer Ther. 2018, 17 (1 Suppl.), B120 DOI: 10.1158/1535-7163.MCT-19-1102

SC239 ADC

SC239 ADC

 

Cancer Res. 2018, 78 (13 Suppl.), 1782 DOI: 10.1158/1538-7445.AM2018-1782

Mal-PEG2-vc-PABC-eribulin ADC

Mal-PEG2-vc-PABC-eribulin ADC

 

Mol. Cancer Ther. 2018, 17, 2665–2675 DOI: 10.1158/1535-7163.MCT-17-1215

MC-Lys-AZ13599185 ADC

MC-Lys-AZ13599185 ADC

 

Cancer Cell. 2016, 29, 117-129 DOI: 10.1016/j.ccell.2015.12.008

NMS249

NMS249

 

Clin. Cancer. Res. 2015, 21, 3298-3306.  DOI: 10.1158/1078-0432.CCR-14-2035

Traceless disulfide-Tubulysin

Traceless disulfide-Tubulysin

 

Nat. Chem. 2016, 8, 1112-1119, DOI : 10.1038/nchem.2635