WO2016115376A1 - Détection et traitement de mélanomes à double résistance aux médicaments - Google Patents

Détection et traitement de mélanomes à double résistance aux médicaments Download PDF

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WO2016115376A1
WO2016115376A1 PCT/US2016/013451 US2016013451W WO2016115376A1 WO 2016115376 A1 WO2016115376 A1 WO 2016115376A1 US 2016013451 W US2016013451 W US 2016013451W WO 2016115376 A1 WO2016115376 A1 WO 2016115376A1
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braf
raf
resistance
brafi
mek1
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Roger S. Lo
Willy HUGO
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The Regents Of The University Of California
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/437—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a five-membered ring having nitrogen as a ring hetero atom, e.g. indolizine, beta-carboline
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4164—1,3-Diazoles
    • A61K31/4184—1,3-Diazoles condensed with carbocyclic rings, e.g. benzimidazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/106—Pharmacogenomics, i.e. genetic variability in individual responses to drugs and drug metabolism
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers

Definitions

  • the present invention relates generally to detection, diagnosis, monitoring and treatment of cancer, such as melanoma.
  • the invention more specifically pertains to cancers resistant to combination therapy using inhibitors of B-RAF and MEK, and selection of effective treatment strategies.
  • RAS and BRAF are frequently mutated in human malignancies.
  • NRAS and, less often, KRAS mutations occur in about 20% of cases and are mutually exclusively with BRAF mutations, which are present in about 50% of cases.
  • Somatic MEK1 or MEK2 mutations which can be concurrent with RAS or BRAF mutations, have also been detected (Hodis et al., 2012; Krauthammer et al., 2012; Nikolaev et al., 2012; Shi et al., 2012a), but their roles in pathogenesis and therapeutic responses remain ill defined.
  • BRAF mutations strongly predict responses to ATP-competitive BRAF inhibitors (BRAFi) such as vemurafenib and dabrafenib.
  • Allosteric MEK1 and MEK2 inhibitors (MEKi) such as trametinib, selumetinib, cobimetinib, and binimetinib, may have antitumor activities against a broader melanoma segment, including those with NRAS mutations or with both wild-type (WT) NRAS and WT BRAF, but MEKi monotherapy for patients with BRAF mutant
  • melanomas is associated with a narrower therapeutic window (versus BRAFi) (Ribas and Flaherty, 2011).
  • NRAS or KRAS mutations Nazarian et al., 2010; Shi et al., 2014), V600E BRAF amplification (Shi et al., 2012b) or alternative splicing (Poulikakos et al., 2011 ; Shi et al., 2012a), MEK1 or MEK2 mutations (Shi et al., 2012a; Wagle et al., 2011), CDKN2A loss (Shi et al., 2014), and genetic alterations in the phosphatidylinositol 3-kinase- phosphatase and tensin homolog-protein kinase B (PI3K-PTEN-AKT) pathway (Shi et al., 2014; Van Allen et al., 2014).
  • PI3K-PTEN-AKT phosphatidylinositol 3-kinase- phosphatase and tensin homolog-protein kinase B
  • intransigence of acquired resistance in response to dual MAPK targeting may be due to preferential emergence of MAPK-redundant resistance pathways.
  • Evidence of branched evolution, extensive interpatient and tumor heterogeneity, and increased tumor fitness as melanoma emerges from BRAFi-imposed evolutionary selection may help explain why the BRAFi+MEKi combinatorial approach is also an "uphill battle" (Shi et al., 2014).
  • the invention provides a method of predicting or detecting the development of acquired resistance to therapeutic effects of combined B-RAF/MEK inhibitor therapy in a patient suffering from cancer.
  • the method comprises assaying a sample obtained from the patient for a measure of combined B-RAF/MEK inhibitor therapy resistance, selecting samples that exhibit a measure of resistance identified in the assaying step; and identifying a patient whose sample was selected in the selecting step as susceptible to developing resistance to combined B-RAF/MEK inhibitor therapy.
  • the measure of resistance is selected from:
  • V600E BRAF amplification concurrent with DUSP4 deletion (9) V600E BRAF amplification concurrent with one or more MEK1 and MEK2 mutations selected from F53Y, Q56P/Q60P, K57N, K59del, V60E, I111S, C121S/C125S, P124IJS G128V, F129L, V154I, E203K, and G276W;
  • Examples of the assaying of the method include, but are not limited to, targeted sequencing, real-time RT-PCR, Sanger sequencing and/or whole exome sequencing.
  • the V600E BRAF ultra-amplification is identified by detecting more than 50 copies, or more than 60 copies, or more than 70 copies of V600E BRAF.
  • ultra- amplification of V600E BRAF is identified by detecting about 74 copies of V600E BRAF.
  • the assaying can comprise, for example, contacting the sample with one or more primers selected from SEQ ID NOs: 1-106.
  • the combined B-RAF/MEK inhibitor therapy comprises treatment with a B-RAF inhibitor selected from vemurafenib and dabrafenib.
  • the combined B-RAF/MEK inhibitor therapy comprises treatment with a MEK inhibitor selected from trametinib, selumetinib, cobimetinib, and binimetinib.
  • the sample is selected from tissue, bodily fluid, blood, tumor biopsy, spinal fluid, and needle aspirate.
  • the method is performed prior to treatment with combined B-RAF/MEK inhibitor therapy.
  • the method is performed after treatment with combined B-RAF/MEK inhibitor therapy.
  • the method is performed during disease progression or clinical relapse on combined B-RAF/MEK inhibitor therapy.
  • the method is performed after suspension of combined B-RAF/MEK inhibitor therapy.
  • the cancer can be any cancer that develops resistance to combined BRAF/MEK inhibitor therapy.
  • the cancer is melanoma.
  • Other cancers include, but are not limited to, lung cancer and other BRAF mutant cancers that are responsive to inhibitors of BRAF, MEK or ERK or their combinations.
  • the method further comprises identifying in a patient who is susceptible to developing or has developed resistance to combined B-RAF/MEK inhibitor therapy the form(s) of resistance and method(s) to counter such resistance.
  • Such methods to counter resistance driven by described mechanisms include (but are not limited to):
  • the method further comprises treating the patient with intermittent dosing of combined B-RAF and MEK inhibitors.
  • the invention additionally provides a method of preventing or suppressing acquired resistance to combined B-RAF/MEK inhibitor therapy in a patient.
  • the method comprises administering to the patient a therapeutic agent selected from the group consisting of:
  • the therapeutic agent is selected from the group consisting of: Omni-RAF, CRAF and RAF paradox breakers, and ERK inhibitors.
  • the therapeutic agent is CCT196969, CCT241161, PLX3397, PLX7904 and/or SCH442984.
  • FIGS 1A-1E Melanomas Resistant to BRAF/MEK Inhibitors Harbor Exaggerated Genetic Mechanisms of BRAF Inhibitor Resistance.
  • DP disease progression on BRAFi; DD-DP, double drug-disease progression.
  • DD-DP melanoma from patient #2 harboring BRAF ultra-amplification by genomic DNA (gDNA) quantitative PCR (Q-PCR; averages of duplicates; two independent primer sets; PMN, peripheral mononuclear cells) and confirmed by Sanger sequencing (baseline tumor, heterozygous V600E allele).
  • gDNA genomic DNA
  • Q-PCR quantitative PCR
  • PMN peripheral mononuclear cells
  • FIGS 2A-2K Melanoma Cells with Acquired BRAFi Resistance Further Resist BRAFi+MEKi by Augmenting Existing or Combining Distinct Mechanisms.
  • BRAFi vemurafenib; MEKi, selumetinib; [inhibitor] range from 0.1 to 2.0 ⁇ at indicated increments.
  • FIGS 3A-3H Melanoma Cells Clonally Develop Resistance to Upfront BRAFi+MEKi via Alternative Genetic Configurations.
  • (3A) Three-day MTT assays (error bars, SEM, n 5; top, relative raw values; bottom, normalized to DMSO vehicle as 100%).
  • the M249 triplet cell lines were plated 16h without inhibitors (BRAFi-vemurafenib; MEKi-selumetinib; ERKi-
  • FIGS 4A-4I Achieving BRAF/MEK Inhibitor Resistance via Tuning V600E BRAF Gene Dosage with or without MEK Mutations.
  • 4A M249 DDR4/5 (plated 16h with BRAFi+MEKi, 1 ⁇ ; transduced with lentiviral shVector or shBRAF for 48h; and treated with the indicated inhibitors at 1 ⁇ for 1h) were analyzed by Western blots (WBs) (TUBULIN, loading control).
  • 4C Cells from B were plated for clonogenic assays.
  • (4G) M249 P engineered to express vector, WT BRAF, or V6OOE/R509 H BRAF (without inhibitors) or M249 DDR4 (BRAFi+MEKi, 1 ⁇ , 16h) were analyzed by WBs.
  • FIGS 5A-5F Distinct MEK Mutants Share Enhanced Interaction with V600E BRAF (5A, 5B, 5C)
  • the M249 triplet cell lines were plated without (P) or with (DDR4/5)
  • BRAFi+MEKi (1 ⁇ , 16h), and lysates were subjected to immunoprecipitation (IP) using the isotype or BRAF- (5A), MEK1- (5B), and MEK2- (5C) specific antibodies.
  • IP immunoprecipitation
  • M249 P engineered to express vector or FLAG- WT MEKI, - F129L MEK1, - Q56P MEK1 concurrent with over-expression of either HA- WT BRAF or HA V600E BRAF were plated with BRAFi+MEKi (1 ⁇ , 16h; except vector control), and the lysates were subjected to IP (anti-lgG or -FLAG). WBs of IP and total fractions. See also Figure 13.
  • FIGS. 6A-6E A BRAF-MEK Interface Critical for V600E BRAF- MUT MEK1 Interaction and Cooperativity in Conferring Double Drug Resistance.
  • Figs. 6A-6B BRAF arginine 662 at a MEK1-BRAF interface.
  • Relative resistance to BRAFi+MEKi assessed over the indicated concentration range and time points. Three repeats (for 0.1 and 1.0 ⁇ ) are shown, and growths were quantified (1 ⁇ ; n 3; normalization relative to V600E BRAF+ MUT M EK1 transduced cells as 100%; means and SD of the mean; *p ⁇ 0.05, ***p ⁇ 0.001 , ns, not significant based on ANOVA). See also Figure 14.
  • FIGS. 7A-7F Resistance to Combined BRAF/MEK Inhibition Results in Exquisite Drug Addiction
  • 7A Clonogenic survival and degrees of drug addiction.
  • Cell were plated (BRAFi+MEKi, 1 ⁇ ; 48 h) and cultured (9 d) with or without specific inhibitor withdrawal (data shown representative of three independent repeats).
  • Figs. 7B, 7C Clonogenic/drug addiction assays as in A except for the indicated high (7B) or low (7C) ERKi (SCH772984) doses starting at 48h.
  • (7D Clonogenic/drug addiction assays comparing SDR vs. DDR cell lines of distinct genetic backgrounds and resistance mechanisms.
  • V600E BRAF melanoma cells can upregulate a V600E BRAF-CRAF-MEK signalsome in response to selection by
  • BRAFi+MEKi This resistansome can consist of a supra-physiologic level of V600E BRAF, which spuriously activate CRAF.
  • V600E BRAF moderately over-expressed V600E BRAF concomitant with a mutant MEK1/2 can lead to increased V600E BRAF- MUT MEK interaction.
  • Both signaling configurations strongly favor ERK activation, leading to growth/survival finely tuned to the BRAFi+MEKi level.
  • WT grey rectangles
  • mutant dark rectangles
  • FIGS 9A-9B Related to Figure 1.
  • the novel F271VPTEN mutation results in loss- of-function.
  • TUBULIN loading control.
  • FIGS 10A-10H Related to Figure 3.
  • vemurafenib vemurafenib
  • MEKi 1 ⁇ selumetinib
  • DMSO vehicle
  • Cellular lysates were collected at the indicated time points after inhibitor treatment and analyzed for the indicated phospho- and total protein levels by Western blotting.
  • TUBULIN loading control.
  • Activation-associated phosphorylation levels of RSK and ERK are dynamically and positively correlated in M249 double drug- resistant sub-lines. Western blot analysis of lysates from M249 isogenic lines for indicated total and phospho-proteins (p-p90RSK Thr573).
  • M249 sub-lines derived to resist a fixed and high BRAFi+MEKi concentration also harbor concomitant BRAF and MEK1 genetic alterations.
  • M249 parental drug-naive cells were treated from the outset with BRAFi (vemurafenib 0.5 ⁇ ) and MEKi (selumetinib 0.5 ⁇ ) every 2-3 days for over three month, and two independent double drug-resistant proliferative subpopulations were isolated and designated as DDR2 and DDR3.
  • Genomic DNA gDNA were isolated from M249 parental and DDR2/3 and analyzed for
  • MEK1 p-values 0.006/0.002 (primer sets 1/2, DDR2 vs. P); 0.09/0.002 (DDR3 vs. P); 0.29/0.59 (DDR4 vs. P); 0.04/0.003 (DDR5 vs. P).
  • Figs. 10F-10G Cellular sensitivities of MAPKi resistance-associated MEK1 mutants to MEKi treatment in the absence of mutant BRAF.
  • FIGS. 11A-11F Related to Figure 4.
  • M395 DDR cells were infected with either shVECTOR or shBRAF lentivirus and cultured in the presence or absence of BRAFi (vemurafenib, 1 ⁇ ) and MEKi (selumetinib, 1 ⁇ ).
  • Western blot analysis showed that BRAF knockdown in the absence of dual inhibitors did not diminish p-ERK levels while in the presence of dual inhibitors BRAF knockdown abrogated both p-CRAF and p-ERK levels.
  • TUBULIN loading control.
  • 11B-11 F V600E BRAF gene dosage gain with or without a MEK1 mutation confers resistance to BRAF/MEK inhibitors but not double-drug addiction.
  • 11 B Western blot analysis of lysates from cell lines from A and B for indicated total and phospho-proteins levels showing the impact of inhibitor pre-conditioning on accelerating p-ERK rebound (parental cell p-ERK rebounds in days). Cells were plated for 24h without inhibitors, treated for 16h (except Parental or Parental+Vector) with BRAF and MEK inhibitors (1 ⁇ ) followed by inhibitor wash-out and continued incubation for 8h.
  • the M249 isogenic triplet cell lines were plated 16h without inhibitors and then exposed to the indicated combinations of inhibitors (BRAFivemurafenib; MEKi- selumetinib). Experiment performed in parallel with that in B.
  • MTT Three-day drug exposure survival assays for the M249 parental cell line engineered with the indicated constructs; the indicated expression of these constructs were initiated by a prior two day withdrawal from doxycycline.
  • FIGS 12A-12C Related to Figure 4. Both V600E BRAF over-expression and MEK1 mutation facilitate resistance to combined BRAF/MEK inhibitors.
  • M395 isogenic cell lines and M395R-engineered cell lines as in A were either treated with DMSO (P) or BRAFi+MEKi (vemurafenib+selumetinib) for 16h or washed free of inhibitors followed by additional 8h of culture. Western blot analysis of given proteins and dynamic p-ERK levels.
  • (12C) Three-day MTT (top) and twelve-day clonogenic (bottom) drug survival assays, using BRAFi alone, MEKi alone, or both BRAFi+MEKi (error bars, SEM, n 5; top, normalized to DMSO vehicle as 100%). At 1 ⁇ of BRAFi+MEKi, in both assays, M395R cell lines engineered to express either MEK1 mutant, survived significantly better than M395R expressing WT MEK1 or M395R (p ⁇ 0.001).
  • FIG. 13 Related to Figure 5. Interaction between wild type and mutant MEK1 and BRAF.
  • the M249 parental cell line was engineered to express vector or FLAG-MEK1 WT, C121S, E203K, K59del, 1111S concurrent with overexpression of either HA- WT BRAF or HA- V600E BRAF.
  • the stable cell lines were then plated with BRAFi+MEKi (1 ⁇ ) for 16h (except vector control) and lysed.
  • the cellular lysates were subjected to immunoprecipitation using the isotype (IgG) or FLAG-specific antibodies.
  • the immunoprecipiated (IP) and total fractions were then probed by Western blots as indicated.
  • Figures 14A-14D The isotype (IgG) or FLAG-specific antibodies.
  • IP immunoprecipiated
  • FIG. 14A Structure of the WT MEKI- WT BRAF heterodimer (KD, kinase domain) (PDB4MNE) showing (i) the face-to-face configuration of the complex, (ii) locations (amino acid residues highlighted with lighter gray near white arrow) of MEK1 residues mutated in melanomas with acquired BRAFi or BRAFi+MEKi resistance, and (iii) locations of BRAF V600 and R509 (i.e., the RAF dimerization interface; thin black arrows).
  • MEK1 residues 1-65 were not resolved in the crystal structure of 4MNE.
  • Figures 15A-15B Related to Figure 7. ERKi-mediated suppression of rebound phospho-ERK unleashed by acute BRAFi and MEKi withdrawal.
  • TGR Tumor growth rates
  • the present invention is based on the discovery of mechanisms of acquired resistance to combined therapy with B-RAF and MEK inhibitors. This discovery enables the following discovery.
  • the invention also provides for implementation of a more effective treatment strategy to manage a specific subset of melanoma patients relapsing on and/or developing resistance to combination therapy.
  • B-RAF inhibitors refers to drugs that target an acquired mutation of B-RAF that is associated with cancer, such as V600E B-RAF.
  • B-RAF inhibitors include PLX4032/vemurafenib or other similar agents, such as
  • V600E -RAF refers to B-RAF having valine (V) substituted for by glutamate (E) at codon 600. Similar nomenclature is used to indicate other amino acid substitutions or deletions ("del").
  • MAK/ERK kinase refers to a mitogen-activated protein kinase also known as microtubule-associated protein kinase (MAPK) or extracellular signal- regulated kinase (ERK).
  • pharmaceutically acceptable carrier includes any material which, when combined with an active ingredient, allows the ingredient to retain biological activity and is non-reactive with the subject's immune system.
  • examples include, but are not limited to, any of the standard pharmaceutical carriers such as a phosphate buffered saline solution, water, emulsions such as oil/water emulsion, and various types of wetting agents.
  • Preferred diluents for aerosol or parenteral administration are phosphate buffered saline or normal (0.9%) saline.
  • compositions comprising such carriers are formulated by well known conventional methods (see, for example, Remington's Pharmaceutical Sciences, 18th edition, A. Gennaro, ed., Mack Publishing Co., Easton, PA, 1990).
  • Methods described herein are performed using clinical samples or biopsies derived from patients or short-term culture derived from same.
  • the methods guide the clinician in stratifying patients for sequential treatment strategies with alternative drug(s) or withdrawal and/or intermittent drug therapy.
  • the invention provides a method of predicting or detecting the development of acquired resistance to therapeutic effects of combined B-RAF/MEK inhibitor therapy in a patient suffering from cancer, the method comprising:
  • V 600E BRAF ultra-amplification e.g., more than 50 copies, or more than 60 copies, or more than 70 copies, such as about 74 copies);
  • kits comprising these primers, or subsets of these primers for targeting mutations identified herein, are also provided by the invention. Such kits may optionally include a nucleotide sequence for use as a positive control in methods of the invention. Also optionally included in the kits are polymerizing enzymes, deoxynucleotide triphosphates (A, G, C, T), and containers for housing each of these components of the kit.
  • the combined B-RAF/MEK inhibitor therapy can comprise, for example, treatment with a B-RAF inhibitor selected from vemurafenib and dabrafenib, and/or treatment with a MEK inhibitor selected from trametinib, selumetinib, cobimetinib, and binimetinib.
  • the sample is selected from tissue, bodily or biologic fluid, such as blood, tumor biopsy, spinal fluid, and needle aspirate.
  • the method can be performed prior to treatment with combined B- RAF/MEK inhibitor therapy, after treatment with combined B-RAF/MEK inhibitor therapy, during disease progression or clinical relapse on combined B-RAF/MEK inhibitor therapy, or after suspension of combined B-RAF/MEK inhibitor therapy.
  • the cancer is melanoma.
  • the method further comprises identifying in a patient who is susceptible to developing or has developed resistance to combined B-RAF/MEK inhibitor therapy the form(s) of resistance and method(s) to counter such resistance.
  • the method can, optionally, further comprise treating the patient with intermittent dosing of combined B-RAF and MEK inhibitors.
  • Methods to counter resistance driven by described mechanisms include (but are not limited to): combinations based on BRAF and MEK inhibitors with the additions of: Inhibitors of the PI3K-AKT-mTORC pathway; Inhibitors downstream of the MAPK pathway such as CDK4/6 inhibitors; Inhibitors that block MEK activation by RAF; Inhibitors that block BRAF and MEK interactions; Inhibitors that block RAF dimerization; and Inhibitors that block RAS functions.
  • inhibitors of the PI3K-AKT-mTORC pathway include, but are not limited to, PI3K p110b inhibitor GSK 2636771 , AKTi GSK690693, and PI3K/mTOR dual inhibitor GSK2126458.
  • inhibitors downstream of the MAPK pathway include, but are not limited to, CDK4/6 inhibitors, e.g., palbocuclib, LEE011 and inhibitors that block MEK activation by RAF.
  • Newer MEK inhibitors target MEK catalytic activity and also impair its reactivation by CRAF, either by dismpting RAF-MEK complexes or by interacting with Ser 222 to prevent MEK phosphorylation by RAF. See Lito P, et al., Cancer Cell. 2014 May 12;25(5):697-710. doi:
  • inhibitors that allosterically block MEK1/2 activities include, but are not limited to, e.g., trametinib, cobimetinib, and selumetinib.
  • Experimental compounds are available that block BRAF and MEK interactions.
  • inhibitors that block RAF dimerization include, but are not limited to, experimental compounds and LY3009120.
  • inhibitors that block RAS functions include, but are not limited to, KRAS G12C inhibitor 6.
  • the invention additionally provides a method of preventing or suppressing acquired resistance to combined B-RAF/MEK inhibitor therapy in a patient.
  • the method comprises administering to the patient a therapeutic agent selected from the group consisting of:
  • the therapeutic agent is selected from the group consisting of: Omni-RAF, CRAF and RAF paradox breakers, and ERK inhibitors.
  • the therapeutic agent is CCT196969, CCT241161, PLX3397, PLX7904 and/or SCH442984.
  • the invention further provides a method of treating a patient having cancer, or who may be at risk of developing cancer or a recurrence of cancer.
  • the patient has melanoma.
  • the melanoma is a B-RAF-mutant melanoma.
  • the cancer can be melanoma or other cancer associated with B-RAF mutation, such as, for example, V600E B-RAF.
  • Patients can be identified as candidates for treatment using the methods described herein. Patients are identified as candidates for treatment on the basis of exhibiting one or more indicators of resistance to B-RAF inhibitor therapy.
  • the treatment protocol can be selected or modified on the basis of which indicators of resistance to B-RAF inhibitor therapy are exhibited by the individual patient.
  • the patient to be treated may have been initially treated with conventional B-RAF inhibitor therapy, or may be a patient about to begin B-RAF inhibitor therapy, as well as patients who have begun or have yet to begin other cancer treatments, including treatment with ERK inhibitors, for example.
  • Patients identified as candidates for treatment with one or more alternative therapies can be monitored so that the treatment plan is modified as needed to optimize efficacy.
  • Examples of alternative therapy include, but are not limited to, intermitting dosing with combined B-RAF/MEK inhibitor therapy, augmenting B-RAF/MEK inhibitor therapy with at least one additional drug.
  • the additional drug can include a MAPK/ERK kinase (MEK) inhibitor, such as PD0325901 , GDC0973, GSK1120212, and/or AZD6244.
  • MEK MAPK/ERK kinase
  • the alternative therapy comprises suspension of vemurafenib therapy.
  • the alternative therapy comprises administering to the patient a MEK inhibitor, optionally in conjunction with vemurafenib therapy, or an inhibitor of the MAPK pathway (RAF, MEK, ERK) in conjunction with an inhibitor of the RTK-PI3K-AKT-mTOR pathway.
  • MEK inhibitors include, but are not limited to PD0325901 , GDC0973, GSK1120212, and/or AZD6244 ⁇ .
  • inhibitors of the RTK-PI3K-AKT-mTOR pathway include, but are not limited to BEZ235, BKM120, PX-866, and GSK2126458.
  • the invention provides a method of preventing or suppressing acquired resistance to combined B-RAF/MEK inhibitor therapy in a patient.
  • the method comprises administering to the patient a therapeutic agent selected from the group consisting of: a suppressor C-RAF kinase activity, and an inhibitor of activation of MEK1 or MEK2 by C- RAF or ERK1 or ERK2 by MEK1 and MEK2.
  • a therapeutic agent include, but are not limited to: Omni-RAF, CRAF and RAF paradox breakers (e.g.,
  • CCT196969, CCT241161 , PLX3397, and PLX7904 CCT196969, CCT241161 , PLX3397, and PLX7904
  • ERK inhibitors e.g., SCH772984
  • the method of preventing or suppressing acquired resistance to combined B- RAF/MEK inhibitor therapy in a patient can be implemented, for example, when such resistance results from MAPK pathway reactivation.
  • MAPK pathway reactivation can be measured by detecting rebound phospho-ERK levels in response to acute BRAF and MEK inhibitor withdrawal or speed of recovery of phospho-ERK levels after a single dose of BRAF and MEK inhibitor treatment.
  • Treatment includes prophylaxis and therapy.
  • Prophylaxis or therapy can be accomplished by a single administration or direct injection, at a single time point or multiple time points to a single or multiple sites. Administration can also be nearly simultaneous to multiple sites.
  • Patients or subjects include mammals, such as human, bovine, equine, canine, feline, porcine, and ovine animals.
  • the subject is preferably a human.
  • treatment comprises administering to a subject a pharmaceutical composition of the invention.
  • a cancer may be diagnosed using criteria generally accepted in the art, including the presence of a malignant tumor.
  • Pharmaceutical compositions may be administered either prior to or following surgical removal of primary tumors and/or treatment such as
  • compositions are administered in any suitable manner, often with
  • Suitable methods of administering treatment in the context of the present invention to a subject are available, and, although more than one route can be used to administer a particular composition, a particular route can often provide a more immediate and more effective reaction than another route.
  • the dose administered to a patient should be sufficient to effect a beneficial therapeutic response in the patient over time, or to inhibit disease progression.
  • the composition is administered to a subject in an amount sufficient to elicit an effective response and/or to alleviate, reduce, cure or at least partially arrest symptoms and/or complications from the disease.
  • An amount adequate to accomplish this is defined as a "therapeutically effective dose.”
  • compositions may be administered, by injection (e.g., intracutaneous, intratumoral, intramuscular, intravenous or subcutaneous), intranasally (e.g., by aspiration) or orally.
  • injection e.g., intracutaneous, intratumoral, intramuscular, intravenous or subcutaneous
  • intranasally e.g., by aspiration
  • between 1 and 10 doses may be administered over a 52 week period.
  • 6 doses are administered, at intervals of 1 month, and booster treatments may be given periodically thereafter.
  • Alternate protocols may be appropriate for individual patients.
  • 2 intradermal injections of the composition are administered 10 days apart.
  • a suitable dose is an amount of a compound that, when administered as described above, is capable of promoting an anti-tumor immune response, and is at least 10-50% above the basal (i.e., untreated) level. Such response can be monitored using conventional methods.
  • the amount of each drug present in a dose ranges from about 100 pg to 5 mg per kg of host, but those skilled in the art will appreciate that specific doses depend on the drug to be administered and are not necessarily limited to this general range.
  • suitable volumes for each administration will vary with the size of the patient.
  • an appropriate dosage and treatment regimen provides the active compound(s) in an amount sufficient to provide therapeutic and/or prophylactic benefit.
  • Such a response can be monitored by establishing an improved clinical outcome (e.g., more frequent remissions, complete or partial, or longer disease-free survival) in treated patients as compared to non-treated patients.
  • Example 1 Tunable-Combinatorial Mechanisms of Acquired Resistance Limit the Efficacy of BRAF/MEK Cotaraetina but Result in Melanoma Drug Addiction
  • Combined BRAF/MEK targeted therapy improves upon BRAF inhibitor (BRAFi) therapy but is still beset by acquired resistance.
  • This Example shows that melanomas acquire resistance to combined BRAF/MEK inhibition by augmenting or combining mechanisms observed in single-agent BRAFi resistance.
  • HiSeq2000 platform were generated. SNVs, insertion-deletions (INDELs), and CNVs were analyzed and visualized as described previously (Shi et al., 2014).
  • BRAF.NRAS, and DUSP4cDNAIevels were quantified by real-timeRT-PCRusing TUBULIN and GAPDH levels for normalization. Relative expressions were calculated using the delta-Ct method.
  • BRAF, NRAS, and DUSP4 gDNA relative copy numbers were quantified by real-time PCR with total gDNA content estimated by assaying the b-globin gene in each sample. All primer sequences are listed in Example 2 below. Sanger sequencing was performed using purified PCR via BigDye v1.1 (Applied Biosystems) in combination with a 3730 DNA Analyzer (Applied Biosystems). WES of M249 triple cell lines were analyzed for shared and distinct genetic alterations and their phylogenetic relationship.
  • PLX4032/vemurafenib (Plexxikon)
  • AZD6244/selumetinib (Selleck Chemicals)
  • SCH772984 (Merck) were made in DMSO.
  • Cell proliferation experiments were performed in a 96-well format (five replicates per sample), drug treatments were initiated 24 hr postseeding for 72 hr, and cell survival was quantified using CellTiter-GLO assay (Promega).
  • Clonogenic assays were performed by plating cells at single-cell density in six-well plates with fresh media and drug replenished every 2 days. Colonies were fixed in 4% paraformaldehyde and stained with 0.05% crystal violet.
  • shBRAF, shCRAF, shPTEN, and shNRAS were subcloned into the lentiviral vector pLL3.7; shDUSP4/pLK0.1 vectors were obtained commercially (Dharmacon).
  • Cell lysates were made in radioimmunoprecipitation assay buffer (Sigma) for direct western blotting or in a PNE buffer (PBS:H20 at 1 : , 0.5% Nonidet P-40, 5mMEDTA, and5% glycerol) for immunoprecipitation, with both buffers supplemented with protease (Roche) and phosphatase (Santa Cruz Biotechnology) inhibitor cocktails.
  • radioimmunoprecipitation assay buffer Sigma
  • PNE buffer PBS:H20 at 1 : , 0.5% Nonidet P-40, 5mMEDTA, and5% glycerol
  • immunoprecipitations were performed using the following antibodies: p-ERKI/2 (T202/Y204), P-MEK1/2 (S217/221), p-AKT (T308), p-CRAF (S338), total ERK1/2, MEK1/2, MEK1 , MEK2, AKT, CRAF, DUSP4, and HA (Cell Signaling Technology); TUBULIN and FLAG (Sigma); BRAF (F-7), BRAF (C-19), p-MEK1 (T291), and p-MEK1 (S222) (Santa Cruz); and p-MEK2 (S226) (United States Biological). Western blot quantification was performed using NIH ImageJ.
  • the 3D structures of MEK1 (3EQC) and PTEN mutants were modeled by the I- TASSER online server. Modeling the V600E BRAF- MUT MEK1 dimer interface was based on the crystal structure of the WT BRAF- WT MEKI dimer (4MNE); the MEK1-KSR2 dimer (2Y4I); and the asymmetric, vemurafenib-bound V600E BRAF dimer (3G07). Protein structures were visualized using PyMol (DeLano Scientific).
  • Supplemental Information providing greater details includes six figures, four tables, and one movie and can be obtained in connection with the publication of this article as Cancer Cell 27, 1-17, February 9, 2015 using dx.doi.org/10.1016/j.ccell.2014.11.018.
  • DD-DP double-drug disease progression
  • n 28 DD-DP tumors, each with patient-matched baseline tumors
  • Figure 1A (1) upfront BRAFi+MEKi (dabrafenib+trametinib or
  • BRAFi+MEKi vemurafenib+cobimetinib
  • DD-DP tumors harboring V600E BRAF amplification four harboring NRAS activating mutations, one harboring a KRAS activating mutation, eight harboring CDKN2A deletions, three harboring PTEN loss-of-function (LOF) mutation (a substitution resulting in F127V; Figure 9) or deletions, and one harboring a PIK3R1 deletion.
  • LEF loss-of-function
  • Dabra, dabrafenib (orally dosed twice a day)
  • Vemu, vemurafenib (orally dosed twice a day)
  • GDC0973 cobimetinib, orally dosed once a day
  • PFS refers to
  • Table 2 Summary of genetic alterations detected in core resistance pathway.
  • BRAFi-Resistant Melanoma Rapidly Upregulates Resistance Mechanisms Individually or Combinatorially to Overcome BRAF/MEK Inhibitors
  • NRAS knockdown restored BRAFi sensitivity to M249 SDR, as would be expected, but it also strongly restored BRAFi+MEKi sensitivity to M249 SDRDDR in both short- and long-term ( Figures 2G and 2H) survival assays, indicating that overexpression of mutant NRAS drove DDR.
  • Figure 2F NRAS knockdown
  • SCH772984 an ERK inhibitor (ERKi) and an analog of which is being tested clinically, was inefficient to inhibit the growth of DDR4 or DDR5 by itself but was highly active against M249 P (Figure 3A).
  • ERKi ERK inhibitor
  • Figure 3A low concentrations of SCH772984 rescued DDR4 and DDR5 from drug addiction, suggesting that suboptimal ERKi dosing to overcome DDR may paradoxically perpetuate DDR fitness.
  • ERKi restored BRAFi+MEKi sensitivity to DDR4 and DDR5, consistent with MAPK pathway reactivation as the major mechanism of acquired resistance to upfront BRAFi+MEKi. This was corroborated by analyzing the MAPK pathway status (p-ERK levels) in the M249 triplet (Figure 3B). After plating for 16 hr without both inhibitors, the triplet cell lines were treated with BRAFi+MEKi (1 hr) at increasing
  • ERKi treatment alone of some melanoma cells previously selected for resistance by BRAFi+MEKi would be ineffective unless very high ERKi doses were delivered, which is unlikely achievable clinically.
  • clonal M249 DDR4 and DDR5 melanoma sublines harbor salient but distinct genetic alterations that represent tunable and combinatorial modes of resistance to BRAFi+MEKi reversible by combining ERKi.
  • V600E BRAF high overexpression induced a robust DDR4-like p-CRAF level
  • V600E BRAF low overexpression concurrent with an MEK1 mutation induced a lower, DDR5-like p-CRAF level.
  • Neither vector control nor MUT MEK1 alone had any impact on the p-CRAF level.
  • supraphysiologic expression of WT BRAF or V600E/R509H braf (known to disrupt BRAF-CRAF dimerization) in M249 P only marginally upregulated p-CRAF (Figure 4G).
  • DDR5 harbored the highest level of D-MEK1 T291, which has been shown to reduce MEK1-MEK2 heterodimerization and MEK2 S226 phosphorylation (Catalanotti et al., 2009) and may also explain the reduced p- MEK1 S222 level ( Figures 5D).
  • V 600E BRAF with BRAF or KSR2 we hypothesized a regulatory V600E BRAF- MUT MEK complex where V600E BRAF R662 makes critical contacts with MEK residues in one complex interface ( Figures 6A and 6B). We predicted that the R662L substitution in V600E BRAF would disrupt this face-to-face V600E BRAF- MUT MEK interaction and attenuate the DDR phenotype. Ectopic expression of vector, HA- WT BRAF, HA- V600E BRAF, and HA- V600E/Ree2L BRAF in WT BRAF
  • HEK293T cells revealed that the R662L substitution did not interfere with the V600E BRAF kinase activation status in the absence of MAPKi ( Figure 6C).
  • M249 P we then engineered M249 P to stably express a FLAG- F129L MEK1 or FLAG- Q60p MEK1 along with HA-tagged WT or various mutant BRAF at levels akin to M249 DDR5 ( Figure 6D).
  • BRAFi+MEKi treatment (1 mM, 16 hr
  • anti-FLAG immunoprecipitation followed by western blots revealed that both MEK1 mutants most abundantly interacted with V600E BRAF, consistent with previous results (Figure 5F).
  • the R662L mutation in the context of V600E BRAF strongly abolished this enhanced V600E BRAF- MUT MEK1 complex and reduced the overall p-ERK levels.
  • V600E/R509HBRAF also appeared to display reduced interaction with MUT MEK1 but without a reduction in the p-ERK levels, suggesting that this apparent reduction was due to loss of BRAF dimers (Figure 6A) (Haling et al., 2014) or higher-order oligomers (Nan et al., 2013) brought down by anti-FLAG. Consistently, whereas engineered M249 P lines highly overexpressing V600E BRAF or minimally overexpressing V600E/R509 H BRAF together with a MEK1 mutant were able to resist robustly BRAFi+MEKi at 1 mM, those cell lines expressing
  • 0n g witn an MEK1 mu tant grew poorly over 28- or 32-day treatments with BRAFi+MEKi ( Figure 6E).
  • Figures 4, 5, and 6; Figures 11-13 highlighted a critical role of upstream MAPK reactivation, i.e., upregulation of the V600E BRAF-CRAF-MEK complex, in the MAPKi resistance phenotype. Buildup of this plastic complex is dependent on the degree of BRAF and/or MEK inhibition and likely other cell context determinants.
  • alternative mechanisms to upregulate this complex can be achieved by V600E BRAF (variably overexpressed) interacting with ⁇ CRAF or with MUTMEK.
  • Table 4 Clinical characteristics of patients followed for melanoma tumor regression or growth deceleration after cessation of MAPK-targeted therapies.
  • V600E BRAF ultra-amplification, G12R NRAS amplification gain-of-function
  • LOF e.g., F127VPTEN, deletions affecting PTEN, CDKN2A, DUSP4
  • SNVs single-nucleotide variants
  • CNVs single-nucleotide variants
  • MEK1 and MEK2 mutants with alterations residing in or proximal to the helices A andC substructures share an increased ability to form an activation-associated complex with V600E BRAF, especially when both BRAF and MEK mutants are moderately overexpressed.
  • a proposed MUTMEK- V600E BRAF heterodimer interface strongly suggests that such a face-to-face physical interaction involves
  • V600E BRAF-CRAF-MEK signaling loop that is highly susceptible to upregulation via single or multiple convergent genetic (and likely nongenetic) alterations.
  • melanoma cell lines with acquired resistance to combined BRAF and MEK inhibition have revealed insights into recent clinical studies. For instance, melanoma cell lines with preexisting BRAFi resistance augment preexisting mechanisms quickly as they adapt to combined BRAF and MEK inhibition. This is consistent with the clinical observation that patients who progressed on BRAFi or MEKi monotherapies infrequently respond to the addition of the other inhibitor, and, for those who do respond sequentially, the responses are generally highly transient. Furthermore, the importance of an MAPKi resistancerelated complex has certain translational implications.
  • Successful strategies targeting this tunable- combinatorial signaling complex may include those inhibiting CRAF function (e.g., omni- or pan-RAF inhibitors), V600E BRAF-CRAF interaction, V600E BRAF-MUTMEK interaction or scaffolding, and MEK activation (e.g., phosphorylation by RAF). These strategies could be built around continued inhibition of mutant BRAF and MEK or alternating regimens.
  • CRAF function e.g., omni- or pan-RAF inhibitors
  • V600E BRAF-CRAF interaction e.g., V600E BRAF-CRAF interaction
  • V600E BRAF-MUTMEK interaction or scaffolding e.g., phosphorylation by RAF

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Abstract

L'invention concerne des méthodes pour prédire ou détecter le développement d'une résistance acquise aux effets thérapeutiques d'un traitement d'association par inhibiteurs de B-RAF/MEK chez un patient atteint de cancer. La méthode consiste à analyser un échantillon prélevé chez un patient pour une mesure de la résistance au traitement d'association par inhibiteurs de B-RAF/MEK, à sélectionner les échantillons qui présentent la mesure de résistance identifiée à l'étape d'analyse ; et identifier un patient dont l'échantillon a été sélectionné à l'étape de sélection comme étant susceptible de développer une résistance au traitement d'association par inhibiteurs de B-RAF/MEK. Des stratégies de traitement peuvent être établies sur la base de la prédiction ou de la détection du développement d'une résistance acquise aux effets thérapeutiques d'un traitement d'association par inhibiteurs de B-RAF/MEK chez un patient atteint de cancer.
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Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2018060415A1 (fr) * 2016-09-29 2018-04-05 Universität Innsbruck Rapporteur de conformation d'activité de kinase pleine longueur
CN108823297A (zh) * 2018-06-13 2018-11-16 领星生物科技（上海）有限公司 基于rt-wes的转录组测序方法
JP2019532051A (ja) * 2016-09-19 2019-11-07 ノバルティス アーゲー Ｒａｆ阻害剤及びｅｒｋ阻害剤を含む治療用組合せ
CN112687354A (zh) * 2019-01-29 2021-04-20 杭州纽安津生物科技有限公司 针对靶向药物PI3K、AKT、mTOR通路的激酶抑制剂的多肽疫苗组合及其设计方法
US11040027B2 (en) 2017-01-17 2021-06-22 Heparegenix Gmbh Protein kinase inhibitors for promoting liver regeneration or reducing or preventing hepatocyte death
WO2021142345A1 (fr) * 2020-01-08 2021-07-15 Icahn School Of Medicine At Mount Sinai Modulateurs à petites molécules de mek liant ksr
WO2022255401A1 (fr) * 2021-06-03 2022-12-08 国立大学法人 東京大学 Marqueur de maladie exprimé en association avec une activation anormale de la voie de erk-mapk
WO2023009572A1 (fr) * 2021-07-27 2023-02-02 Verastem, Inc. Polythérapie pour le traitement d'une croissance cellulaire anormale
US11873296B2 (en) 2022-06-07 2024-01-16 Verastem, Inc. Solid forms of a dual RAF/MEK inhibitor
US12036227B2 (en) 2017-05-02 2024-07-16 Novartis Ag Combination therapy
WO2024186926A1 (fr) * 2023-03-06 2024-09-12 The Regents Of The University Of California Stratégies pour empêcher l'évolution d'un mélanome cutané létal largement métastatique résistant aux polythérapies
US12187703B2 (en) 2019-05-13 2025-01-07 Novartis Ag Crystalline forms of N-(3-(2-(2-hydroxyethoxy)-6-morpholinopyridin-4-yl)-4-methvlphenyl)-2 (trifluoromethyl)isonicotinamide as Raf inhibitors for the treatment of cancer

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20130217721A1 (en) * 2010-11-19 2013-08-22 The Regents Of The University Of California Compositions and methods for detection and treatment of b-raf inhibitor-resistant melanomas

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20130217721A1 (en) * 2010-11-19 2013-08-22 The Regents Of The University Of California Compositions and methods for detection and treatment of b-raf inhibitor-resistant melanomas

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
LIU ET AL.: "Activities of multiple cancer-related pathways are associated with BRAF mutation and predict the resistance to BRAF/MEK inhibitors in melanoma cells", CELL CYCLE, vol. 13, no. Issue 2, 2014, pages 208 - 219 *
LONG ET AL.: "Increased MAPK reactivation in early resistance to dabrafenib /trametinib combination therapy of BRAF-mutant metastatic melanoma", NATURE COMMUNICATIONS, vol. 5, no. 5694, 2014, pages 1 - 9 *
MONTAGUT ET AL.: "Elevated CRAF as a potential mechanism of acquired resistance to BRAF inhibition in melanoma", CANCER RESEARCH, vol. 68, no. 12, 2008, pages 4853 - 4861 *
MORICEAU ET AL.: "Tunable-combinatorial mechanisms of acquired resistance limit the efficacy of BRAF/MEK cotargeting but result in melanoma drug addiction", CANCER CELL, vol. 27, 15 January 2015 (2015-01-15), pages 240 - 256 *
VILLANUEVA ET AL.: "Concurrent MEK2 mutation and BRAF amplification confer resistance to BRAF and MEK inhibitors in melanoma", CELL REPORTS, vol. 4, 2013, pages 1090 - 1099 *

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US12011449B2 (en) 2016-09-19 2024-06-18 Novartis Ag Therapeutic combinations comprising a c-RAF inhibitor
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JP7114575B2 (ja) 2016-09-19 2022-08-08 ノバルティス アーゲー Ｒａｆ阻害剤及びｅｒｋ阻害剤を含む治療用組合せ
US12130292B2 (en) 2016-09-29 2024-10-29 Kincon Biolabs Gmbh Full length kinase activity-conformation reporter
WO2018060415A1 (fr) * 2016-09-29 2018-04-05 Universität Innsbruck Rapporteur de conformation d'activité de kinase pleine longueur
US11237173B2 (en) 2016-09-29 2022-02-01 Universitat Innsbruck Full length kinase activity-conformation reporter
US11040027B2 (en) 2017-01-17 2021-06-22 Heparegenix Gmbh Protein kinase inhibitors for promoting liver regeneration or reducing or preventing hepatocyte death
US12036227B2 (en) 2017-05-02 2024-07-16 Novartis Ag Combination therapy
CN108823297A (zh) * 2018-06-13 2018-11-16 领星生物科技（上海）有限公司 基于rt-wes的转录组测序方法
CN112687354B (zh) * 2019-01-29 2022-07-12 杭州纽安津生物科技有限公司 针对靶向药物PI3K、AKT、mTOR通路的激酶抑制剂的多肽疫苗组合及其设计方法
CN112687354A (zh) * 2019-01-29 2021-04-20 杭州纽安津生物科技有限公司 针对靶向药物PI3K、AKT、mTOR通路的激酶抑制剂的多肽疫苗组合及其设计方法
US12187703B2 (en) 2019-05-13 2025-01-07 Novartis Ag Crystalline forms of N-(3-(2-(2-hydroxyethoxy)-6-morpholinopyridin-4-yl)-4-methvlphenyl)-2 (trifluoromethyl)isonicotinamide as Raf inhibitors for the treatment of cancer
WO2021142345A1 (fr) * 2020-01-08 2021-07-15 Icahn School Of Medicine At Mount Sinai Modulateurs à petites molécules de mek liant ksr
WO2022255401A1 (fr) * 2021-06-03 2022-12-08 国立大学法人 東京大学 Marqueur de maladie exprimé en association avec une activation anormale de la voie de erk-mapk
WO2023009572A1 (fr) * 2021-07-27 2023-02-02 Verastem, Inc. Polythérapie pour le traitement d'une croissance cellulaire anormale
US11873296B2 (en) 2022-06-07 2024-01-16 Verastem, Inc. Solid forms of a dual RAF/MEK inhibitor
WO2024186926A1 (fr) * 2023-03-06 2024-09-12 The Regents Of The University Of California Stratégies pour empêcher l'évolution d'un mélanome cutané létal largement métastatique résistant aux polythérapies

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