Fig.1 Detection of CRBN expression in spleen, brain, liver, intestine and muscle tissues by RT-PCR.?

Wild type: only one band at 185 bp with primers F1/R1 (mCrbn);

Homozygous: one band at 262 bp with primers F2/R2 (hCRBN) and one band at 374 bp with primers F3/R3 (hCRBN);

Abbr. M, DNA marker; HO, homozygous;? WT, wild type.

Fig.2 Expression characterization of hCRBN in homozygous hCRBN KI mice by Western blot.

Fig. 3 The degradation of GSPT1, CK1a and Myc by Lenalidomide and CC-885 in hCRBN knockin mice were determined by western blot.

The splenocytes derived from wild-type C57BL/6 (WT) and homozygous hCRBN KI (HO) mice were treated with 1% DMSO, Lenalidomide (10 nM, 100 nM, 1000 nM) and CC-885 (10 nM, 100 nM, 1000 nM) ,respectively, in RPMI-1640 cell culture medium for 18 hours ex vivo.

Fig.4 Blood concentration curves of Lenalidomidein in C57BL/6 and HO hCRBN mice (female, 10wks, n=3).

（In cooperation with a client）

Fig.5 Blood concentration of CC-885 in C57BL/6 and? hCRBN mice 1h after first administration (female, 10wks). (In cooperation with a client）

Fig.6 Mean body weight (A) and survival curve (B) of C57BL/6 and HO hCRBN KI mice treated with CC-885 or vehicle (n=8/group).?

All hCRBN mice treated with CC-885 dead within 2 days, while mice from other groups survived. The result indicates that CC-885 exhibits significant toxicity only in hCRBN humanized mice (In cooperation with a client).

Fig.7 The degradation of GSPT1, CK1a and Myc by CC-885 in C57BL/6 and hom hCRBN knockin mice in vivo (In cooperation with a client).

The wild-type C57BL/6 (WT, group 1&2) and homozygous hCRBN KI (HO, group 3&4) mice were treated with vehicle (group 1&3)? and 5 mg/kg CC-885 (group 2&4) , respectively. The mice were anesthetized, perfused, and tissues were harvested at 6 hr post the 2nd administration.

Fig.8 Partial representative pictures of small intestine pathology (Magnification: 200x).

(In cooperation with a client)

Fig.9 Partial representative pictures of liver pathology (Magnification: 200x).

(In cooperation with a client)

Fig.10 Partial representative pictures of spleen pathology (Magnification: 200x).

(In cooperation with a client)