WO2018223112A1 - Prédiction des résultats d'un traitement du cancer de l'estomac - Google Patents

Prédiction des résultats d'un traitement du cancer de l'estomac Download PDF

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WO2018223112A1
WO2018223112A1 PCT/US2018/035788 US2018035788W WO2018223112A1 WO 2018223112 A1 WO2018223112 A1 WO 2018223112A1 US 2018035788 W US2018035788 W US 2018035788W WO 2018223112 A1 WO2018223112 A1 WO 2018223112A1
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tubb3
tymp
peptide
docetaxel
cisplatin
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PCT/US2018/035788
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Todd Hembrough
Fabiola CECCHI
Sarit SCHWARTZ
Christina Yau
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Expression Pathology, Inc.
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Priority to EP18809143.3A priority Critical patent/EP3630187A1/fr
Priority to CN201880035577.2A priority patent/CN110678203A/zh
Priority to AU2018275156A priority patent/AU2018275156A1/en
Priority to JP2019566297A priority patent/JP2020522500A/ja
Priority to KR1020197036676A priority patent/KR20200012895A/ko
Priority to CA3065333A priority patent/CA3065333A1/fr
Publication of WO2018223112A1 publication Critical patent/WO2018223112A1/fr

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    • A—HUMAN NECESSITIES
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    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/337—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having four-membered rings, e.g. taxol
    • A—HUMAN NECESSITIES
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    • A61K31/00—Medicinal preparations containing organic active ingredients
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    • 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
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
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    • A61K31/00—Medicinal preparations containing organic active ingredients
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    • 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
    • A61K31/513—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim having oxo groups directly attached to the heterocyclic ring, e.g. cytosine
    • 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
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    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/10—Transferases (2.)
    • C12N9/1048—Glycosyltransferases (2.4)
    • C12N9/1077—Pentosyltransferases (2.4.2)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y204/00—Glycosyltransferases (2.4)
    • C12Y204/02—Pentosyltransferases (2.4.2)
    • C12Y204/02004—Thymidine phosphorylase (2.4.2.4)
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
    • G01N33/57407—Specifically defined cancers
    • G01N33/57446—Specifically defined cancers of stomach or intestine
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6803—General methods of protein analysis not limited to specific proteins or families of proteins
    • G01N33/6848—Methods of protein analysis involving mass spectrometry
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2300/00—Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/52—Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis

Definitions

  • Treatment 1 comprising administration of 5-fluoruracil/folinic acid (DeGramont) versus a regimen (treatment 2) comprising a first administration of the FOLFIRI regimen
  • Treatment 1 comprises 5FU/LV while Treatment 2 comprises FOLFIRI + docetaxel/cisplatin.
  • SRJV1 mass spectrometry is used to measure TUBB3 and TYMP proteins in tumor cells derived from patient tumor tissue and these measurements are then used to identify cancer patients, and in particular gastric cancer patients, most likely to respond to treatment with the sequential administration of FOLFIRI followed by combination docetaxel/cisplatin chemotherapy agents.
  • TUBB3 also known as class III ⁇ -tubulin, is a critical component of the cell microtubule assembly andis also actively involved in regulating ligand binding. Proteomic analysis has revealed that many factors bound to these cysteine residues are involved in the oxidative stress and glucose deprivation response. TUBB3 has been investigated as both a prognostic biomarker and an indicator of resistance to docetaxel and other similar taxane compounds where reports implicate high quantitative levels of TUBB3 as a biomarker of poor outcome. Accordingly, determining quantitative expression levels of the TUBB3 protein in patient cancer cells could help determine how cancer cells will respond to treatment with docetaxel.
  • TYMP also known as thymidine phosphorylase, is a protein that synthesizes dTMP from thymine and is part of the dTMP biosynthesis salvage pathway, which is itself part of pyrimidine metabolism. TYMP is known as an angiogenic factor which promotes angiogenesis in vivo and stimulates the in vitro growth of a variety of endothelial cells.
  • TYMP normally has a highly restricted target cell specificity, acting only on endothelial cells. In some cases, however, TYMP can be abnormally expressed at high levels in tumor cells. TYMP functions to convert 5-dFUR to 5FU and 5FU to FdUM,P which promotes the functionality of 5FU to inhibit the TS protein, thereby blocking the production of DNA and inducing tumor cell apoptosis.
  • TUBB3 and TYMP are prognostic predictors of therapeutic outcome in cancer patients and as such can provide information about chemotherapy treatment strategies of cancer.
  • the presence and/or quantitative levels of TUBB3 and TYMP protein expression in patient tumor cells procured from patient tumor tissue is determined by quantitating a specified peptide derived from subsequences of each of the TUBB3 and TYMP full-length proteins using the methodology of SRM mass spectrometry.
  • Specific quantitative levels of the TUBB3 protein as detected by SRM mass spectrometry in cancer cells present within a cancer patient indicates that the patient is either more likely or less likely to respond in a positive manner to a chemotherapy regimen containing docetaxel.
  • Specific quantitative levels of the TYMP protein as detected in cancer cells within a cancer patient by SRM mass spectrometry indicates that the patient is more likely or less likely to respond in a positive manner to a chemotherapy regimen that includes 5FU.
  • Irinotecan also known as camptosar
  • camptosar is a cancer chemotherapy agent that interacts with DNA by intercalation, thereby inhibiting the progression of the enzyme topoisomerase I (TOPOl) which relaxes supercoils in DNA for transcription.
  • TOPOl enzyme topoisomerase I
  • Doxorubicin stabilizes the TOPOl complex after it has broken the DNA chain for replication, preventing the DNA double helix from being resealed and stopping the process of replication. This prevents cancer cells from synthesizing DNA and stops cancer cell division and tumor growth.
  • TOPOl enzyme in cancer cells can overcome the effects of irinotecan and cause resistance to the effects of irinotecan in the cancer cells, allowing them to synthesize DNA and promoting cellular division and tumor growth.
  • Fluorouracil also known as adrucil, is a chemotherapy agent that functions by blocking DNA, thereby inhibiting cell division and preventing tumor cells from dividing and growing.
  • 5-FU acts in several ways, but acts principally as a thymidylate synthase (TS) inhibitor. Interrupting TS activity blocks synthesis of the pyrimidine thymidine, which is a nucleoside required for DNA replication. Thymidylate synthase methylates deoxyuridine monophosphate (dUMP) to form thymidine monophosphate (dTMP).
  • dUMP deoxyuridine monophosphate
  • dTMP thymidine monophosphate
  • Administration of 5-FU causes a scarcity in dTMP, and rapidly dividing cancerous cells undergo cell death due to this lack of thymine.
  • High levels of thymidylate synthase can overcome the effects of 5-FU while high levels of the thymidine phosphorylase (TYMP) protein promotes the activity of
  • Docetaxel also known as taxotere, is a semi-synthetic analogue of paclitaxel (Taxol). It is in the drug class of taxanes. Docetaxel binds to microtubules reversibly with high affinity, stabilizing microtubules and preventing depolymerization, thus killing dividing cells. This stabilization of the cellular microtubule assembly leads to a significant decrease in free tubulin (TUBB3), needed for microtubule formation, and results in inhibition of mitotic cell division between metaphase and anaphase, preventing further cancer cell division and tumor growth. High expression levels of the TUBB3 protein in cancer cells can overcome the effects of docetaxel and thus provide resistance to the effects of docetaxel in the cells, allowing them to depolymerize microtubule assembly and thus promote cellular division and tumor growth.
  • TUBB3 free tubulin
  • Cisplatin also known as platinol, is a cancer chemotherapy agent that interferes with DNA replication thus preventing cells from dividing and leading to tumor cell death via apoptosis. Cisplatin irreversibly crosslinks DNA in several different ways, interfering with mitotic cell division. The damaged DNA elicits DNA repair mechanisms, which in turn activate apoptosis when repair proves impossible, thus killing the tumor cells.
  • Leucovorin calcium also known as folinic acid, does not have cancer-fighting properties but is a medication used to decrease the toxic side effects of chemotherapeutic agents. LV stabilizes binding of fluorodeoxy uridine monophosphate (FdUMP) to thymidylate synthase by increasing the intracellular pool of 5,10-methylene tetrahydrofolate (CH2THF), and thus enhancing thymidylate synthase (TS) inhibition).
  • FdUMP fluorodeoxy uridine monophosphate
  • CH2THF 5,10-methylene tetrahydrofolate
  • TS thymidylate synthase
  • Leucovorin has almost no side effects of its own but when used in combination with fluorouracil it can increase the severity of side effects of that drug.
  • Figure 1 shows the Kaplan Meier overall survival (OS) curves using TUBB3 ⁇ 750amol ⁇ g and TUBB3 > 750amol ⁇ g cutoff for this gastric cancer cohort treated with treatment 1 regimen comprising 5FU/LV.
  • OS overall survival
  • Figure 2 shows the Kaplan Meier overall survival (OS) curves using TYMP ⁇ 1335amol ⁇ g and TYMP > 1335amol ⁇ g cutoff for this gastric cancer cohort treated with treatment 1 regimen comprising 5FU/LV.
  • OS overall survival
  • Figure 3 shows the Kaplan Meier overall survival (OS) curves using TUBB3 ⁇ 750amol ⁇ g and TUBB3 > 750amol ⁇ g cutoff for this gastric cancer cohort treated with treatment 2 regimen comprising FOLFIRI followed subsequently by administration of the combination docetaxel/cisplatin.
  • OS overall survival
  • Figure 4 shows the Kaplan Meier overall survival (OS) curves using TYMP ⁇
  • Figure 5 shows the Kaplan Meier overall survival (OS) curves using TYMP ⁇ 2800amol ⁇ g and TYMP > 2800amol ⁇ g cutoff for this gastric cancer cohort treated with treatment 2 regimen comprising FOLFIRI followed subsequently by administration of the combination docetaxel/cisplatin.
  • OS overall survival
  • Figure 6 shows the Kaplan Meier overall survival (OS) curves using the combination of TUBB3 cutoff of 750amol ⁇ g + TYMP cutoff of 1300amol ⁇ g for this gastric cancer cohort treated with treatment 2 regimen comprising FOLFIRI followed subsequently by administration of the combination docetaxel/cisplatin.
  • OS overall survival
  • Figure 7 shows the Kaplan Meier overall survival (OS) curves using the combination of TUBB3 cutoff of 750amol ⁇ g + TYMP cutoff of 2800amol ⁇ g for this gastric cancer cohort treated with treatment 2 regimen comprising FOLFIRI followed subsequently by administration of the combination docetaxel/cisplatin.
  • OS overall survival
  • Methods are provided for determining if a cancer patient, and specifically a gastric patient, will clinically respond in a favorable manner to the therapeutic strategy comprising a first administration of the FOLFIRI regimen (irinotecan/5-fluoruracil/folinic acid) followed by a separate sequential administration of the combination of the chemotherapy agents docetaxel and cisplatin (FOLFIRI + docetaxel/cisplatin).
  • FOLFIRI inotecan/5-fluoruracil/folinic acid
  • diagnostic methods for measuring the TUBB3 and TYMP proteins in a tumor sample or samples from the patient are provided.
  • the sample is advantageously formalin-fixed.
  • an SRM/MRM assay that simultaneously measures a specific TUBB3 peptide fragment and a specific TYMP peptide fragment, and particular characteristics about the peptide fragments, the amount of the FOLFIRI regimen
  • TUBB3 and TYMP proteins in cells derived from formalin fixed paraffin embedded (FFPE) tissue is determined.
  • the peptide fragments derive from the full-length TUBB3 and TYMP proteins, wherein the peptide sequence for TUBB3 protein is SEQ ID NO: l
  • this SRM/MRM assay can measure these peptides directly in complex protein lysate samples prepared from cells procured from patient tissue samples, such as formalin fixed cancer patient tissue.
  • patient tissue samples such as formalin fixed cancer patient tissue.
  • Methods of preparing protein samples from formalin-fixed tissue are described in U.S. Pat. No. 7,473,532, the contents of which are hereby incorporated by reference in their entirety.
  • the methods described in U.S. Pat. No. 7,473,532 may conveniently be carried out using Liquid Tissue reagents and protocol available from Expression Pathology Inc. (Rockville, Md.).
  • formalin fixed, paraffin embedded tissue The most widely and advantageously available form of tissue, and cancer tissue, from cancer patients is formalin fixed, paraffin embedded tissue. Formaldehyde/formalin fixation of surgically removed tissue is by far and away the most common method of preserving cancer tissue samples worldwide and is the accepted convention in standard pathology practice.
  • Aqueous solutions of formaldehyde are referred to as formalin. "100%" formalin consists of a saturated solution of formaldehyde (this is about 40% by volume or 37% by mass) in water, with a small amount of stabilizer, usually methanol, to limit oxidation and degree of polymerization.
  • Results from the SRM/MRM assay can be used to correlate accurate and precise quantitative levels of the TUBB3 and TYMP proteins within the specific cancer of the patient from whom the tissue was collected and preserved, including gastric cancer tissue. This not only provides diagnostic/prognostic information about the cancer, but also permits a physician or other medical professional to determine appropriate therapy for the patient. In this case, utilizing these assays can provide information about specific levels of TUBB3 and TYMP protein expression simultaneously in cancer tissue and whether or not the patient from whom the cancer tissue was obtained will respond in a favorable way to the therapeutic strategy comprising administering the FOLFIRI regimen (irinotecan/5-fluoruracil/folinic acid) followed by a separate sequential administration of the combination of the FOLFIRI regimen (irinotecan/5-fluoruracil/folinic acid) followed by a separate sequential administration of the combination of the FOLFIRI regimen (irinotecan/5-fluoruracil/folinic acid) followed by a separate sequential administration of the combination of the FOLFI
  • chemotherapy agents docetaxel and cisplatin (FOLFIRI + docetaxel/cisplatin).
  • Treating cancer patients with the FOLFIRI regimen is a common and effective strategy that has been shown to prolong the lives of cancer patients, especially gastric cancer patients.
  • One of the agents in the combination therapy FOLFIRI regimen is 5-fluoruracil (5FU) which functions in multiple ways that involve blocking the action of thymidylate synthase and thus stopping the production of DNA as well as blocking the production of RNA and thus causing apoptosis.
  • the TYMP protein is involved in converting 5-dFUR to active 5FU as well as converting 5FU to FUDR. Both of these conversions promote the activity of exogenously administered 5FU and thus enhance tumor cell killing by 5FU and higher levels of the TYMP protein are desirable when treating the cancer patient with a regimen that includes 5FU.
  • Treating cancer patients with docetaxel is also a common and effective strategy for prolonging the lives of the patients.
  • the TUBB3 protein functions as a critical component of the cell microtubule assembly and is also actively involved in regulating ligand binding. Both functions are involved in helping cells to grow and divide. Docetaxel inhibits normal microtubule processes and ligand binding and thus prevent tumor cells from growing and dividing. It therefore is useful for a clinician to know quantitative levels of the TUBB3 protein in a patient's cancer cells because the therapeutic effects of docetaxel in those cells can be overcome simply be overexpression of the TUBB3 protein. If a clinician knows that a cancer patient's tumor cells express very high levels of TUBB3 protein he/she will likely not prescribe docetaxel to the patient because the patient is unlikely to respond favorably.
  • the clinician will be more likely to prescribe docetaxel as one of the chemotherapy agents in a combination therapy strategy because the drug will likely be able to inhibit normal microtubule function and ligand binding in the cancer cells and thus help to shut down growth of the tumor.
  • IHC immunohistochemistry
  • Inaccurate IHC test results may mean that patients diagnosed with cancer do not receive the best possible care. If all or part of a cancer is positive for a specific target oncoprotein but test results classify it as negative, physicians are unlikely to recommend the correct therapeutic treatment, even though the patient could potentially benefit from those agents. If a cancer is oncoprotein target negative but test results classify it as positive, physicians may recommend a specific therapeutic treatment, even though the patient is unlikely to get any benefits and is exposed to the agent's secondary risks. Thus there is great clinical value in the ability to correctly evaluate quantitative levels of the TUBB3 and TYMP proteins in tumors, especially gastric tumors, so that the patient will have the greatest chance of receiving the most optimal treatment.
  • Detection of peptides and determining quantitative levels of specified TUBB3 and TYMP fragment peptides are determined in a mass spectrometer by the SRM/MRM methodology, whereby the SRM/MRM signature chromatographic peak area of each peptide is determined within a complex peptide mixture present in a Liquid Tissue lysate (see U. S. Pat. No. 7,473,532, as described above).
  • Quantitative levels of the TUBB3 and TYMP proteins are then determined by the SRM/MRM methodology whereby the SRM/MRM signature chromatographic peak area of an individual specified peptide from each of the TUBB3 and TYMP proteins in one biological sample is compared to the SRM/MRM signature chromatographic peak area of a known amount of a "spiked" internal standard for each of the individual specified TUBB3 and TYMP fragment peptides.
  • the internal standard is a synthetic version of the same exact TUBB3 and TYMP fragment peptides where the synthetic peptides contain one or more amino acid residues labeled with one or more heavy isotopes.
  • Such isotope labeled internal standards are synthesized so that mass spectrometry analysis generates a predictable and consistent SRM/MRM signature chromatographic peak that is different and distinct from the native TUBB3 and TYMP fragment peptide chromatographic signature peaks and which can be used as comparator peaks.
  • the SRM/MRM signature chromatographic peak area of the native peptide is compared to the SRM/MRM signature chromatographic peak area of the internal standard peptide, and this numerical comparison indicates either the absolute molarity and/or absolute weight of the native peptide present in the original protein preparation from the biological sample.
  • Quantitative data for fragment peptides are displayed according to the amount of protein analyzed per sample.
  • the mass spectrometer In order to develop the SRM/MRM assay for the TUBB3 and TYMP fragment peptides additional information beyond simply the peptide sequence may be utilized by the mass spectrometer. That additional information is important in directing and instructing the mass spectrometer, (e.g., a triple quadrupole mass spectrometer) to perform the correct and focused analysis of the specified TUBB3 and TYMP fragment peptides.
  • An important consideration when conducting an SRM/MRM assay is that such an assay may be effectively performed on a triple quadrupole mass spectrometer.
  • That type of a mass spectrometer may be considered to be the most suitable instrument for analyzing a single isolated target peptide within a very complex protein lysate that may consist of hundreds of thousands to millions of individual peptides from all the proteins contained within a cell.
  • the additional information provides the triple quadrupole mass spectrometer with the correct directives to allow analysis of a single isolated target peptide within a very complex protein lysate that may consist of hundreds of thousands to millions of individual peptides from all the proteins contained within a cell.
  • SRM/MRM assays can be developed and performed on any type of mass spectrometer, including a MALDI, ion trap, ion trap/quadrupole hybrid, or triple quadrupole, presently the most advantageous instrument platform for SRM/MRM assay is often considered to be a triple quadrupole instrument platform.
  • the additional information about target peptides in general, and in particular about the specified TUBB3 and TYMP fragment peptides, may include one or more of the mono isotopic mass of each peptide, its precursor charge state, the precursor m/z value, the m/z transition ions, and the ion type of each transition ion.
  • the peptide sequences of the specified TUBB3 and TYMP fragment peptides are shown in Table 1.
  • tumor samples are obtained from a cohort of patients suffering from cancer, for example gastric cancer.
  • the tumor samples are formalin-fixed using standard methods and the level of TUBB3 and TYMP in the samples is measured using the methods as described above.
  • the tissue samples may also be examined using IHC and FISH using methods that are well known in the art.
  • the patients in the cohort are treated with the combination of either: 1) the chemotherapy strategy (treatment 1) comprising administration of 5-fluoruracil/folinic acid (DeGramont); or 2) the chemotherapy treatment strategy (treatment 2) comprising a first administration of the FOLFIRI regimen (irinotecan/5-fluoruracil/folinic acid) followed by a sequential administration of combination docetaxel/cisplatin.
  • Treatment 1 comprises 5FU/LV while Treatment 2 comprises FOLFIRI + docetaxel/cisplatin.
  • Patient response is measured using methods that are well known in the art, for example by recording the overall survival of the patients at time intervals after treatment.
  • a suitable reference level can be determined using statistical methods that are well known in the art, for example by determining the lowest p value of a log rank test.
  • a reference level can be used to identify those patients whose TUBB3 and TYMP protein expression levels indicate that they may likely benefit from the combination of treatment regimen 1 or treatment regimen 2.
  • the FOLFIRI regimen comprising irinotecan + 5-fluoruracil + folinic acid is a common treatment regimen for gastric cancer patients.
  • Levels of TUBB3 and TYMP proteins in patient tumor samples typically are expressed in amol ⁇ g, although other units can be used.
  • a reference level can be expressed as a range around a central value, for example, +/- 250, 150, 100, 50 or 25 amol/ ⁇ g.
  • both nucleic acids and protein can be analyzed from the same Liquid Tissue biomolecular preparation it is possible to generate additional information about disease diagnosis and drug treatment decisions from the nucleic acids in the same sample in which proteins were analyzed. For example, if the TUBB3 and TYMP proteins are expressed by certain cells at increased levels, when assayed by SRM the data can provide information about the state of the cells and their potential for uncontrolled growth, choice of optimal therapy, and potential drug resistance. At the same time, information about the status of genes and/or the nucleic acids and proteins they encode (e.g., mRNA molecules and their expression levels or splice variations) can be obtained from nucleic acids present in the same Liquid Tissue biomolecular preparation. Nucleic acids can be assessed simultaneously with the SRM analysis of proteins, including the TUBB3 and TYMP proteins. In one
  • information about the TUBB3 and TYMP proteins and/or one, two, three, four or more additional proteins may be assessed by examining the nucleic acids encoding those proteins.
  • Those nucleic acids can be examined, for example, by one or more, two or more, or three or more of: sequencing methods, polymerase chain reaction methods, restriction fragment polymorphism analysis, identification of deletions, insertions, and/or determinations of the presence of mutations, including but not limited to, single base pair polymorphisms, transitions, transversions, or combinations thereof.
  • TUBB3 was assessed by determining the association between its protein levels and OS in a univariate Cox model for treatment 1 regimen comprising, 5FU/LV.
  • a cutoff threshold was derived for dichotomizing patients by TUBB3 levels, using Cox proportional modeling and Gehan- Breslow-Wilcoxon significance test. Half the patients were randomly selected, balancing for the number of OS events and cohort (first vs. second), as the training set. Every possible value between the 25th and 75th percentile was evaluated as a potential threshold to dichotomize the training set into 'High' vs.
  • the median OS of the TUBB3 ⁇ 750amol ⁇ g group is 1325 days; while the median OS of the TUBB3 > 750amol ⁇ g group is 1991 days. The results are shown in Figure 1.
  • 57 patients had TYMP levels ⁇ 1335amol ⁇ g and 65 patients had TYMP levels >1335amol ⁇ g.
  • Patients with TYMP levels > 1335amol ⁇ g had significantly better OS than those with TYMP levels ⁇ 1335amol ⁇ g.
  • Figure 4 shows the association between TYMP and OS in the gastric cancer patient population treated with treatment regimen 2 comprising FOLFIRI (irinotecan/5- fluoruracil/folinic acid) followed subsequently by administration of the combination docetaxel/cisplatin.
  • FOLFIRI irinotecan/5- fluoruracil/folinic acid
  • the previously derived cutoff of 1335amol ⁇ g was selected for comparison to the earlier analysis.

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Abstract

L'invention concerne des méthodes visant à identifier si un patient atteint de cancer, et en particulier un patient atteint d'un cancer de l'estomac, répondra à un traitement consistant en une stratégie thérapeutique séquentielle comprenant une première administration du protocole FOLFIRI (irinotécan/5-fluoruracile/acide folinique) suivie d'une administration séquentielle séparée de l'association des agents chimiothérapeutiques docétaxel et cisplatine (FOLFIRI + docétaxel/cisplatine). Les fragments peptidiques TUBB3 et TYMP spécifiés sont détectés et quantifiés avec précision par spectrométrie de masse en mode SRM directement dans des cellules tumorales, et en particulier des cellules tumorales du cancer de l'estomac, qui sont collectées à partir de tissu tumoral provenant d'un patient atteint de cancer, et les mesures sont comparées à des niveaux de référence afin de déterminer si les patients atteints de cancer répondront positivement à un traitement par le traitement d'association séquentiel FOLFIRI + docétaxel/cisplatine.
PCT/US2018/035788 2017-06-02 2018-06-04 Prédiction des résultats d'un traitement du cancer de l'estomac WO2018223112A1 (fr)

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EP3510409A4 (fr) * 2016-09-07 2020-03-25 Expression Pathology, Inc. Dosage srm/mrm pour la protéine de la chaîne bêta -3 de la tubuline (tubb3)

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LU ET AL.: "Expressions of Thymidylate Synthase, Thymidine Phosphorylase, Class III beta- tubulin, and Excision Repair Cross complementing Group 1 predict Response in Advanced Gastric Cancer Patients Receiving Capecitabine Plus Paclitaxel or Cisplatjn", CHIN J CANCER RES., vol. 23, no. 4, December 2011 (2011-12-01), pages 288 - 294, XP019991207 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP3510409A4 (fr) * 2016-09-07 2020-03-25 Expression Pathology, Inc. Dosage srm/mrm pour la protéine de la chaîne bêta -3 de la tubuline (tubb3)

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