Inhibitors BRAF (gene)

figure 3: b-raf kinase domain locked in inactive conformation bound bay43-9006. hydrophobic interactions anchor bay43-9006 in atp binding site while urea group hydrogen-bonding traps d594 of dfg motif. bay43-9006 trifluoromethyl phenyl ring further prohibits dfg motif , activation loop movement active confermer via steric blockage.

bay43-9006 (sorafenib, nexavar)is v600e mutant b-raf , c-raf inhibitor approved fda treatment of primary liver , kidney cancer. bay43-9006 disables b-raf kinase domain locking enzyme in inactive form. inhibitor accomplishes blocking atp binding pocket through high-affinity kinase domain. binds key activation loop , dfg motif residues stop movement of activation loop , dfg motif active conformation. finally, trifluoromethyl phenyl moiety sterically blocks dfg motif , activation loop active conformation site, making impossible kinase domain shift conformation become active.

the distal pyridyl ring of bay43-9006 anchors in hydrophobic nucleotide-binding pocket of kinase n-lobe, interacting w531, f583, , f595. hydrophobic interactions catalytic loop f583 , dfg motif f595 stabilize inactive conformation of these structures, decreasing likelihood of enzyme activation. further hydrophobic interaction of k483, l514, , t529 center phenyl ring increase affinity of kinase domain inhibitor. hydrophobic interaction of f595 center ring decreases energetic favorability of dfg conformation switch further. finally, polar interactions of bay43-9006 kinase domain continue trend of increasing enzyme affinity inhibitor , stabilizing dfg residues in inactive conformation. e501 , c532 hydrogen bond urea , pyridyl groups of inhibitor respectively while urea carbonyl accepts hydrogen bond d594 s backbone amide nitrogen lock dfg motif in place.

the trifluoromethyl phenyl moiety cements thermodynamic favorability of inactive conformation when kinase domain bound bay43-9006 sterically blocking hydrophobic pocket between αc , αe helices dfg motif , activation loop inhabit upon shifting locations in active conformation of protein.

vemurafenib

figure 4: structures of vemurafenib (right) , precursor, plx 4720 (left), 2 inhibitors of active conformation of b-raf kinase domain

plx4032 (vemurafenib) v600 mutant b-raf inhibitor approved fda treatment of late-stage melanoma. unlike bay43-9006, inhibits inactive form of kinase domain, vemurafenib inhibits active dfg-in form of kinase, firmly anchoring in atp-binding site. inhibiting active form of kinase, vemurafenib selectively inhibits proliferation of cells unregulated b-raf, cause cancer.

since vemurafenib differs precursor, plx4720, in phenyl ring added pharmacokinetic reasons, plx4720 s mode of action equivalent vemurafenib s. plx4720 has affinity atp binding site partially because anchor region, 7-azaindole bicyclic, differs natural adenine occupies site in 2 places nitrogen atoms have been replaced carbon. enables strong intermolecular interactions n7 hydrogen bonding c532 , n1 hydrogen bonding q530 preserved. excellent fit within atp-binding hydrophobic pocket (c532, w531, t529, l514, a481) increases binding affinity well. ketone linker hydrogen bonding water , difluoro-phenyl fit in second hydrophobic pocket (a481, v482, k483, v471, i527, t529, l514, , f583) contribute exceptionally high binding affinity overall. selective binding active raf accomplished terminal propyl group binds raf-selective pocket created shift of αc helix. selectivity active conformation of kinase further increased ph-sensitive deprotonated sulfonamide group stabilized hydrogen bonding backbone peptide nh of d594 in active state. in inactive state, inhibitor s sulfonamide group interacts backbone carbonyl of residue instead, creating repulsion. thus, vemurafenib binds preferentially active state of b-raf s kinase domain.