WO2017175111A1

WO2017175111A1 - Strn-alk fusion as a therapeutic target in colorectal cancer

Info

Publication number: WO2017175111A1
Application number: PCT/IB2017/051890
Authority: WO
Inventors: Eric SLOSBERG; Barinder KANG; August SALVADO; Steven STEIN
Original assignee: Novartis AG
Current assignee: Novartis AG
Priority date: 2016-04-04
Filing date: 2017-04-03
Publication date: 2017-10-12
Anticipated expiration: 2018-10-04

Abstract

The present disclosure relates to an ALK inhibitor for use in treating colorectal cancer, characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide; to an ALK inhibitor for use in a method of selectively treating a patient having colorectal cancer; to a kit for use in predicting the likelihood that a patient having a colorectal cancer will respond to treatment with an ALK inhibitor; and to a kit for use in treating a patient having colorectal cancer.

Description

STRN-ALK FUSION AS A THERAPEUTIC TARGET IN COLORECTAL

CANCER

FIELD OF THE DISCLOSURE

The present disclosure relates to an ALK inhibitor for use in treating a mammalian cancer, particularly colorectal cancer in human; use of an ALK inhibitor for the preparation of a medicament for the treatment of colorectal cancer; methods of selectively treating a patient having colorectal cancer with an ALK inhibitor or a drug other than ALK inhibitor; a method of or a kit for use in predicting the likelihood that a patient having a colorectal cancer will respond to treatment with an ALK inhibitor; a kit for use in treating a patient having colorectal cancer and other closely related methods and kits.

BACKGROUND OF THE DISCLOSURE

Colorectal cancer (CRC) is a major cause of cancer-related deaths worldwide. Today, the clinical management of patients with initially unresectable metastatic CRC (mCRC) primarily consists of chemotherapy. However, because existing therapies can be toxic, more specific therapeutic regimens such as targeted agents have been sought to improve the outcomes and quality of life of patients with colorectal cancer. Identification of essential tumor growth drivers has initiated a new era of targeted cancer therapy. Such targeted therapies as the epidermal growth factor receptor (EGFR)-targeting monoclonal antibodies (mAbs) cetuximab and panitumumab and the vascular endothelial growth factor (VEGF)-targeting mAb bevacizumab led to a marked improvement in clinical outcome for patients with mCRC.

Efforts to identify alterations that could predict benefit from a targeted therapy approach in colorectal cancer have proved difficult. Up to date, only limited number of gene alterations has been detected in colorectal cancer. For example, KRAS mutation analysis is an accepted molecular approach in colorectal cancer. However, unlike the demonstration of EGFR mutation or ALK rearrangement in non-small cell lung cancer (NSCLC), which are used to select patients for targeted therapies, KRAS mutational status is instead used to exclude patients unlikely to benefit from monoclonal anti-EGFR therapy.

There is a continuing need in the art for uncovering so far unrecognized subsets of colorectal cancer that may harbor genetic alterations which would help us predict response to targeted therapies.

SUMMARY OF THE DISCLOSURE

It is an object of the present disclosure to provide a novel STRN-ALK gene fusion as a targetable genomic alteration in colorectal cancer, and to provide an ALK inhibitor for use in treating colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide.

In accordance with the present disclosure a novel STRN-ALK gene fusion was surprisingly identified as a targetable genomic alteration by comprehensive genomic profiling in a patient with advanced metastatic colorectal cancer, who failed standard therapy. The patient was subsequently treated with ALK targeted tyrosine kinase inhibitor ceritinib achieving a dramatic clinical response. Present disclosure provides an evidence that patients with colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide benefit from the treatment with an ALK inhibitor. Knowledge of the fact that the STRN-ALK fusion can drive the proliferation of colorectal cancer and that the cancer can respond to an ALK inhibitor can be applied, among others, when characterizing a sample of a colorectal cancer, or for providing means to detect said alteration in the colorectal cancer, such as for example a method for detecting the STRN-ALK gene fusion or a STRN-ALK fusion polypeptide in a biological sample or a kit comprising a probe that is capable of detecting the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide.

In one aspect, the present disclosure relates to an ALK inhibitor for use in treating colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN- ALK fusion polypeptide.

In another aspect, the disclosure provides the use of an ALK inhibitor for the preparation of a medicament for the treatment of colorectal cancer, wherein the colorectal cancer is characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide.

In another aspect, the present disclosure relates to a method of selectively treating a patient having colorectal cancer with an ALK inhibitor, wherein the method comprises the steps of:

(a) selecting the patient for the treatment with an ALK inhibitor on the basis of the patient having the colorectal cancer characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide; and

(b) thereafter, administering a therapeutically effective amount of an ALK inhibitor to the patient.

In yet another aspect, the present disclosure relates to a method of selectively treating a patient having colorectal cancer, comprising either:

(a) selectively administering a therapeutically effective amount of an ALK inhibitor to the patient on the basis of the patient having the colorectal cancer characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide; or (b) selectively administering a therapeutically effective amount of a drug other than an ALK inhibitor to the patient on the basis of the patient not having the colorectal cancer characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide.

In a further aspect, the present disclosure relates to a method of selectively treating a patient having colorectal cancer with an ALK inhibitor, comprising:

(a) assaying a biological sample obtained from the patient for a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide;

(b) thereafter, selecting the patient for the treatment with an ALK inhibitor on the basis of the patient having the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide; and

(c) thereafter, administering a therapeutically effective amount of an ALK inhibitor to the patient.

In yet a further aspect, the present disclosure relates to a method of selectively treating a patient having colorectal cancer, comprising:

(a) assaying a biological sample obtained from the patient for STRN-ALK gene fusion or a STRN-ALK fusion polypeptide; and

(b) thereafter, selectively administering to the patient either:

(i) a therapeutically effective amount of an ALK inhibitor on the basis of the biological sample obtained from the patient having the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide; or

(ii) a therapeutically effective amount of a drug other than an ALK inhibitor on the basis of the biological sample obtained from the patient not having the STRN- ALK gene fusion or the STRN-ALK fusion polypeptide. In another aspect, the present disclosure relates to a method of predicting the likelihood that a patient having a colorectal cancer will respond to treatment with an ALK inhibitor, comprising assaying a biological sample obtained from the patient for the presence or absence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide, wherein:

(a) the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide is indicative of an increased likelihood that the patient will respond to treatment with an ALK inhibitor; and

(b) the absence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide is indicative of a decreased likelihood that the patient will respond to treatment with an ALK inhibitor. In a further aspect, the present disclosure relates to a kit for use in predicting the likelihood that a patient having colorectal cancer will respond to the treatment with an ALK inhibitor comprising:

(a) at least one probe capable of detecting the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide; and

(b) instructions for using the probe to assay a biological sample from the colorectal cancer patient for the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide, wherein the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide is indicative of an increased likelihood that the patient will respond to treatment with an ALK inhibitor and the absence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide is indicative of a decreased likelihood that the patient will respond to treatment with an ALK inhibitor.

In yet a further aspect, the present disclosure relates to a kit for use in treating a patient having colorectal cancer comprising:

(a) a therapeutically effective amount of an ALK inhibitor;

(b) at least one probe capable of detecting the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide;

(c) instructions for using the probe to assay a biological sample obtained from the patient for the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide,

(d) instructions for administering the ALK inhibitor to the patient if the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide is present in the biological sample obtained from the patient ; and

(e) optionally, means for administering the ALK inhibitor to the patient.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 Histologic Analysis of the Tumor. Gross image of a 15.1 cm cecal tumor (panel A). A Representative histologic section of the tumor shows moderately differentiated adenocancer containing glands exhibiting prominent infoldings, pseudopapillary structures, and a prominent mucinous component (Panel B, hematoxylin and eosin).

FIG. 2 Molecular Analysis of the Tumor. Screenshot of Integrative Genomics Viewer demonstrating uniquely mapped paired-end reads in the intronic regions of STRN (Intron 3) and ALK (Intron 19) on chromosome 2 (Panel C). Schematic representation of the STRN-ALK fusion of the N-terminal portion of STRN and C-terminal portion of ALK. Exons 1-3 of STRN containing the caveolin-binding domain (CB) and coiled-coil domain (CC) are shown in blue. Exons 20-29 of ALK containing the tyrosine kinase domain (TK) are shown in orange. Both fusion partners are located on the short arm of chromosome 2 (2p22.2 and 2p23, separated by ~ 7.69 Mb), indicating that the fusion is a result of intrachromosomal paracentric rearrangement. The coiled-coil STRN domain may act as a dimerization motif that could constitutively activate ALK tyrosine kinase (Panel D).

FIG. 3 Immunohistochemical Analysis of the Tumor. Immunohistochemistry with the D5F3 anti-ALK antibody showing strong diffuse intracytoplasmic immunoreactivity confirming ALK overexpression as a result of STRN-ALK fusion. The loss of ALK extracellular and intramembrane domains detected by sequencing is associated with intracellular localization of the fusion protein (Panel E). Dual-color break-apart fluorescence in situ hybridization (FISH) analysis of the tumor cells performed with a 5' ALK probe (green) and 3' ALK probe (red) (panel F). Single isolated red probe signals (arrow) indicate the ALK chromosomal rearrangement. An unsplit red and green probe signals indicate the non-rearranged wild type ALK locus (arrowhead).

FIG. 4 Umbilical Skin Metastasis and Imaging Studies before and after Ceritinib Treatment. A pretreatment photograph shows the tumor metastasis involving periumbilical skin (Panel A). Skin metastasis significantly decreased within 7 weeks of treatment with ceritinib (Panel B). After an additional 2 weeks of treatment there is marked decrease in periumbilical thickening (Panel C) At 6 months follow-up the mass protruding through the skin resolved, leaving a scar (Panel D). Pretreatment CT scan shows large pelvic tumor mass (Panel E). Six months after ceritinib therapy a CT scan demonstrates resolution of all contrast-enhancing tumor (panel F).

FIG. 5 Schematic representation of the STRN-ALK fusion of the N-terminal portion of STRN and C-terminal portion of ALK.

DETAILED DESCRIPTION OF THE DISCLOSURE

Use of specific therapeutic regimens is more beneficial for a patient, as it is proven to be less toxic and more effective. Previous studies uncovered only limited subset of colorectal cancer characterized by specific genetic signatures, which allow predicting response to targeted therapies. The inventors have now identified a transforming STRN- ALK fusion as a therapeutic target in colorectal cancer, and illustrated the potential for personalized molecular based therapy for the treatment of advanced malignancies. The disclosure is based on the identification of a novel transforming ALK fusion event involving STRN gene in colorectal cancer patients. Advantageously, the disclosure can be used for predicting the likelihood that a patient having a colorectal cancer will respond to treatment with an ALK inhibitor based on the presence or absence of STRN-ALK gene fusion, and thus specifically select patients having colorectal cancer who will benefit from treatment with an ALK inhibitor. In one aspect, the present disclosure relates to an ALK inhibitor for use in treating colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN- ALK fusion polypeptide.

An ALK inhibitor can also be used in treating colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide that has progressed or is intolerant to chemotherapy. Similarly, the ALK inhibitor can be used in treating colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide that has progressed or is intolerant to epidermal growth factor receptor (EGFR) inhibitor. A physician evaluates if the disease is stable and would know how to measure cancer progression. Equally, a physician would be able to recognize adverse effects of the first line therapy, such as chemotherapy or EGFR inhibitor, if necessary, after discussion with the patient.

The terms "a" and "an" and "the" and similar references in the context of describing the disclosure (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Where the plural form is used for compounds, patients, cancers and the like, this is taken to mean also a single compound, patient, or the like.

The term "ALK", as used herein, refers to anaplastic lymphoma kinase, also known as ALK tyrosine kinase receptor or CD246 (cluster of differentiation 246). The Entrez Gene ID is 238.

The term "STRN", as used herein, refers to striatin (calmodulin binding protein). The Entrez Gene ID is 6801. STRN encodes a calcium-dependent calmodulin-binding protein.

The term "STRN-ALK gene fusion", as used herein, refers to abnormal DNA rearrangement where the STRN gene is fused to the ALK gene. This abnormal gene fusion leads to the production of a fusion protein (STRN-ALK), referred herein as "STRN-ALK fusion polypeptide". As both fusion partners are located on the short arm of chromosome 2 (2p22.2 and 2p23, separated by ~ 7.69 Mb), the fusion is a result of intrachromosomal paracentric rearrangement. For example, as disclosed herein, an STRN-ALK gene fusion may involve the intrachromosomal translocation of exons 1-3 of STRN to exons 20-29 of ALK (Fig. 2, panel D). The predicted fusion protein resulting from this gene fusion does not have a calmodulin-binding domain and the WD-repeats, but retains the N-terminal caveolin-binding and the coiled-coil binding domains of STRN, fused to the intracellular juxtamembrane region of ALK, containing the kinase domain (Fig. 2, panel D and Fig. 5). The STRN-ALK fusion protein comprises amino acids residues 1 - 137 of STRN portion and 1059 - 1620 of ALK portion (Fig. 2, panel D and Fig. 5). The fusion between the fusion partners is between amino acid 137 of STRN and amino acid 1059 of ALK. The coiled-coil STRN domain may act as a dimerization motif that could constitutively activate ALK tyrosine kinase. In the present disclosure, an ALK inhibitor can be a compound that inhibits ALK with the IC50 of less than 100 μΜ, preferably less than 10 μΜ, more preferably less than ΙμΜ, measured by a Caliper mobility shift assay. The Caliper mobility shift technology is based on the separation of particles of different charges and sizes in an electrical field, similar to capillary electrophoresis. The Caliper kinase assays utilize fluorescently labeled peptides as kinase substrates. The phosphorylation of the peptide in the course of the reaction introduces additional negative charges via the phosphate and hence permits its separation from the phosphorylated peptide. Both, the separation and the detection of the labeled peptides take place in the microfluidic system of the Caliper Lab Chip. The LabChips have 12 "sippers" enabling the parallel analysis of 12 samples at the same time. The fact that both, unphosphorylated peptide (substrate) and phosphorylated peptide (product) are measured and that the separation makes the readout relatively insensitive to interference by fluorescent compounds results in the excellent data quality of this assay. General assay procedure can be performed at 30°C for 60 min in a total volume of 9 μΐ_^ including 0.050 μΐ_^ of compound dilution or pure DMSO, respectively. The reaction can be terminated by the addition of 16 of stop solution (100 mM Hepes, 5 % (v/v) DMSO, 0.1 % (v/v) Coating reagent, 10 mM EDTA, 0.015 % (v/v) Brij 35). After termination of the reactions, the plates are transferred into the Caliper LabChip 3000 workstation for analysis. The effect of a compound on the enzymatic activity is obtained from the linear progress curves in the absence and presence of the compound and routinely determined from one reading (end point measurement).

According to the present disclosure, the ALK inhibitor can be for example a compound selected from the group consisting of

AE684 Alectiniij rizoti.nlb

AP26113 A5P3026 AZD3463 and 5-chloro-N2-(2-isopropoxy-5-methyl-4-(piperidin-4-yl)phenyl)-N4-[2-(propane-2- sulfonyl)-phenyl]-pyrimidine-2,4-diamine, or a pharmaceutically acceptable salt thereof.

The term "pharmaceutically acceptable salts" refers to salts that retain the biological effectiveness and properties of the compound when used according to this disclosure and, which typically are not biologically or otherwise undesirable. Pharmaceutically acceptable acid addition salts can be formed with inorganic acids and organic acids, e.g., acetate, aspartate, benzoate, besylate, bromide / hydrobromide, bicarbonate / carbonate, bisulfate/sulfate, camphorsulfonate, chloride/hydrochloride, chlortheophyllonate, citrate, ethandisulfonate, fumarate, gluceptate, gluconate, glucuronate, oleate, oxalate, palmitate, pamoate, phosphate/hydrogen phosphate/dihydrogen phosphate, propionate, stearate, succinate, subsalicylate, tartrate, tosylate, trifluoroacetate salt or the like. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like.

In a preferred embodiment, the present disclosure relates to the ALK inhibitor 5- chloro-N2-(2-isopropoxy-5-methyl-4-(piperidin-4-yl)phenyl)-N4-[2-(propane-2- sulfonyl)-phenyl]-pyrimidine-2,4-diamine, or a pharmaceutically acceptable salt thereof. The compound 5-chloro-N2-(2-isopropoxy-5-methyl-4-(piperidin-4-yl)phenyl)-N4-[2- (propane-2-sulfonyl)-phenyl]-pyrimidine-2,4-diamine, also known under name ceritinib, is a compound of formula I, and is described in Example 7 (Compound 66) of

WO2008/073687.

The term "colorectal cancer", as used herein, refers to cancer in the colon or rectum, also known as colon cancer, rectal cancer or bowel cancer. In one embodiment, the present disclosure relates to metastatic colorectal cancer.

The term "treatment" as used herein comprises a treatment relieving, reducing or alleviating at least one symptom in a subject, increasing progression-free survival, overall survival, extending duration of response or delaying progression of a disease. For example, treatment can be the diminishment of one or several symptoms of a disorder or complete eradication of a disorder, such as cancer. Within the meaning of the present disclosure, the term "treatment" also denotes to arrest, delay the onset (i.e., the period prior to clinical manifestation of a disease) and/or reduce the risk of developing or worsening a disease in a patient, e.g., a mammal, particularly the patient is a human. The term "treatment" as used herein comprises an inhibition of the growth of a tumor incorporating a direct inhibition of a primary tumor growth and / or the systemic inhibition of metastatic cancer cells.

An ALK inhibitor can be used in treating colorectal cancer characterized by the presence a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide, wherein the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide has been detected in a biological sample from the cancer obtained from a patient having said cancer.

The term "biological sample", as used herein, refers to a biological specimen taken by sampling so as to be representative of any other specimen taken from the source of the specimen. In one embodiment, a biological sample is cells or tissue from the cancer obtained from a patient having said cancer.

A "subject," "individual" or "patient" is used interchangeably herein, which refers to a vertebrate, preferably a mammal, more preferably a human. Mammals include, but are not limited to, mice, simians, humans, farm animals, sport animals, and pets.

Before treatment, a patient population can be stratified according to the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide. For example, a patient having colorectal cancer can be first selected for the treatment with an ALK inhibitor on the basis of the patient having STRN-ALK gene fusion or the STRN-ALK fusion polypeptide; and thereafter, a therapeutically effective amount of an ALK inhibitor is administered to the patient. Patient can be selected based on the presence of the fusion. Depending on the outcome of the selection step, or the characterization of the colorectal cancer that a patient has, the patient can be administered an ALK inhibitor. For example, a patient can be administered a therapeutically effective dose of the ALK inhibitor once it has been determined that the fusion is present in the cancer. In alternative, if it turns out that the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide are not present in the patient sample or a cancer sample , the patient can be administered some other drug which may be better suited to treat the special cancer subtype.

The presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide can also be used as an indicator of an increased likelihood that the patient will respond to treatment with an ALK inhibitor. Therefore, the result of detection step can tell a physician or an informed person whether a patient is likely going to respond to the treatment with an ALK inhibitor. Generally, the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide can be indicative of an increased likelihood that the patient will respond to treatment with an ALK inhibitor; and the absence of the STRN- ALK gene fusion or the STRN-ALK fusion polypeptide can be indicative of a decreased likelihood that the patient will respond to treatment with an ALK inhibitor. As used herein, "selecting" and "selected" in reference to a patient is used to mean that a particular patient is specifically chosen from a larger group of patients on the basis of (due to) the particular patient having a predetermined criteria. Similarly, "selectively treating" refers to providing treatment to a patient having a particular disease, where that patient is specifically chosen from a larger group of patients on the basis of the particular patient having a predetermined criteria. Similarly, "selectively administering" refers to administering a drug to a patient that is specifically chosen from a larger group of patients on the basis of (due to) the particular patient having a predetermined criteria. By selecting, selectively treating and selectively administering, it is meant that a patient is delivered a personalized therapy based on the patient's particular biology, rather than being delivered a standard treatment regimen based solely on the patient having a particular disease. Selecting, in reference to a method of treatment as used herein, does not refer to fortuitous treatment of a patient that has the biomarker, but rather refers to the deliberate choice to administer treatment to a patient based on the patient having the biomarker. Thus, selective treatment differs from standard treatment, which delivers a particular drug to all patients, regardless of their biomarker.

The term "a therapeutically effective amount" of a compound of the present disclosure refers to an amount of the compound of the present disclosure that will elicit the biological or medical response of a subject, for example, reduction or inhibition of an enzyme or a protein activity, or ameliorate symptoms, alleviate conditions, slow or delay disease progression, or prevent a disease, etc.

A therapeutically effective amount of an ALK inhibitor in vivo may range depending on the route of administration, between about 0.05 to about 50 mg per kg body weight per day, preferably about 0.1-25 mg/kg/day, more preferably from about 0.5-10 mg/kg/day, in single or divided doses. For a 70 kg human, this would amount to a preferable dosage range of about 35-700 mg per day. Daily dose of ceritinib can be for example 750 mg. The recommended dose and schedule for crizotinib is 250 mg orally, twice daily, with or without food. Alectinib can be for example used by administering 300 mg twice daily.

The patient having colorectal cancer can be selected for the treatment with an ALK inhibitor based on assaying a biological sample obtained from the patient for STR -ALK gene fusion or the STRN-ALK fusion polypeptide. An ALK inhibitor can be then used in treating a patient having colorectal cancer characterized in that:

(a) a biological sample obtained from the patient is assayed for STRN-ALK gene fusion or the STRN-ALK fusion polypeptide; and

a therapeutically effective amount of an ALK inhibitor is selectively administered to the patient on the basis of the biological sample from the patient having STRN-ALK gene fusion or the STRN-ALK fusion polypeptide. The term "assaying" is used to refer to the act of identifying, screening, probing or determining, which act may be performed by any conventional means. For example, a sample may be assayed for the presence of a particular marker by using an ELISA assay, a Northern blot, imaging, etc. to detect whether that marker is present in the sample. The terms "assaying" and "determining" contemplate a transformation of matter, e.g., a transformation of a biological sample, e.g., a blood sample or other tissue sample, from one state to another by means of subjecting that sample to physical testing. Further, as used herein, the terms "assaying" and "determining" are used to mean testing and/or measuring. The phrase "assaying a biological sample from the patient for..." and the like is used to mean that a sample may be tested (either directly or indirectly) for either the presence or absence of a given factor or for the level of a particular factor. It will be understood that, in a situation where the presence of a substance denotes one probability and the absence of a substance denotes a different probability, then either the presence or the absence of such substance may be used to guide a therapeutic decision.

The assaying or detection for STRN-ALK gene fusion or the STRN-ALK fusion polypeptide can comprise a technique selected from the group consisting of Next Generation Sequencing (NGS), Northern blot analysis, polymerase chain reaction (PCR), reverse transcription-polymerase chain reaction (RT-PCR), TaqMan-based assays, direct sequencing, dynamic allele-specific hybridization, high-density oligonucleotide SNP arrays, restriction fragment length polymorphism (RFLP) assays, primer extension assays, oligonucleotide ligase assays, Southern Blot, immunoassays, immunohistochemistry, ELISA, fluorescence in situ hybridization analysis (FISH), kinase activity assays, flow cytometry, Western blot, HPLC, and mass spectrometry. In a preferred embodiment, the step of assaying or detection for STRN-ALK gene fusion comprises PCR, RT-PCR or fluorescence in situ hybridization analysis (FISH). In another preferred embodiment, the step of assaying or detection for the STRN-ALK fusion polypeptide comprises immunohistochemistry. Techniques are generally known to a person skilled in biochemistry, microbiology and diagnostic tests. In addition, the techniques can be applied according to explanation and instruction found in further references such as for example WO2008/127248, WO2012162373 or references cited therein.

An ALK inhibitor can be administered for at least 6 months. In a preferred embodiment, a therapeutically effective amount of the ALK inhibitor is administered for at least 6 months. In a further embodiment, the ALK inhibitor leads to a progression free survival of at least 6 months.

The ALK inhibitor can be administered as a first line or as a second line treatment.

Generally, until further clinical data is generated, the ALK inhibitor is administered after a patient has been given a standard of care, but has relapsed or has become intolerant to the standard of care, or the cancer has become resistant to the first line medicaments. However, it is expected that targeted therapy may provide clinical benefit over the standard of care and thus it can be valuable to administer the ALK inhibitor as a first line treatment. Therefore, the present disclosure also relates to the administration of the ALK inhibitor as a first line treatment. An ALK inhibitor can also be used for the preparation of a medicament for the treatment of colorectal cancer, wherein the colorectal cancer is characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide.

As used herein, "predicting" indicates that the methods described herein provide information to enable a health care provider to determine the likelihood that an individual having the disorder will respond to or will respond more favorably to treatment. It does not refer to the ability to predict response with 100% accuracy. Instead, the skilled artisan will understand that it refers to an increased probability.

As used herein, "likelihood" and "likely" is a measurement of how probable an event is to occur. It may be used interchangeably with "probability". Likelihood refers to a probability that is more than speculation, but less than certainty. Thus, an event is likely if a reasonable person using common sense, training or experience concludes that, given the circumstances, an event is probable. In some embodiments, once likelihood has been ascertained, the patient may be treated (or treatment continued, or treatment proceed with a dosage increase) with the test compound. In one embodiment, the "likelihood" and "likely" denote a chance in percent of how probable an event is to occur.

The phrase "increased likelihood" refers to an increase in the probability that an event will occur. For example, some methods herein allow prediction of whether a patient will display an increased likelihood of responding to treatment with the test molecule or an increased likelihood of responding better to treatment with the test molecule. In one embodiment the increased likelihood means that there is more than 50% chance, more than 60 % chance, more than 70 % or more than 80 % chance that an event will occur. Equally, a decreased likelihood means, that the chance is lower than 50%, lower than 60 %, lower than 70 % or lover than 80 %, respectively, that an event will occur.

The diagnosis or the determination of whether a patient with a colorectal cancer can be done by using a kit for use in predicting the likelihood that a patient having colorectal cancer will respond to the treatment with an ALK inhibitor comprising:

In one embodiment, the probe is an oligonucleotide that specifically hybridizes to a region of a nucleic acid coding for the STRN-ALK gene fusion, or an oligonucleotide that specifically hybridizes to a region of a nucleic acid coding for an equivalent genetic marker of the STR -ALK gene fusion, or the probe is an antibody that binds to the STRN-ALK fusion polypeptide. Further details on how to prepare for example an antibody that binds to a specific region of a polypeptide can be found in literature such as in WO2008/127248, WO2012162373 or references cited therein. Generally, well known techniques can be used in preparing, testing and validating a suitable probe that binds to a STRN-ALK gene fusion. Similarly, a process of preparing and selecting, testing and validating an antibody that binds to a specific target is also well known in the art. In the same manner the antibody can be prepared that binds to the STRN-ALK fusion polypeptide

The probe can be an oligonucleotide that specifically hybridizes to a region of a nucleic acid coding for the STRN-ALK gene fusion, or an oligonucleotide that specifically hybridizes to a region of a nucleic acid coding for an equivalent genetic marker of the STRN-ALK gene fusion, or the probe is an antibody that binds to the STRN-ALK fusion polypeptide.

The following Examples illustrates the disclosure described above, but is not, however, intended to limit the scope of the disclosure in any way. Other test models known as such to the person skilled in the pertinent art can also determine the beneficial effects of the claimed disclosure.

Examples

Example 1:

SUMMARY:

A novel STRN-ALK gene fusion was identified as a targetable genomic alteration by comprehensive genomic profiling utilizing the FoundationOne (Foundation Medicine, Cambridge, MA) test platform in a patient with advanced metastatic colorectal cancer who failed standard therapy. The patient was treated with ALK targeted tyrosine kinase inhibitor ceritinib achieving a dramatic clinical response. This case provides evidence that patients with colorectal cancer driven by ALK tyrosine kinase may benefit from specific targeted monotherapy.

METHODS:

Case Report:

The patient is an 87-year old woman without significant past medical history who presented three years ago with fatigue, hemoglobin of 8.6 gm/dL and iron deficiency. Colonoscopy revealed a cecal mass and the patient underwent a right hemicolectomy. Pathologic examination of the specimen revealed a 5.1 cm tumor that penetrated to the visceral peritoneal surface and involved 7 of 28 lymph nodes (Fig. 1, panel A). Upon review of the histology, the tumor exhibited features of a moderately differentiated adenocarcinoma with glands showing infolding, cribriforming, pseudopapillary structures and a prominent mucinous component (Fig. 1, panel B). Analysis of mismatch repair proteins by immunohistochemistry revealed loss of expression of MLH1 and PMS2 with retained expression of MSH2 and MSH6. Staging chest computed tomography (CT) revealed small bilateral pulmonary nodules suspicious for metastatic disease.

The patient received 10 cycles of mFOLFOX6 with resolution of radiographic abnormalities. She was without evidence of disease until she presented with peritoneal carcinomatosis 18 months later. She received 8 treatments of FOLFIRI then received maintenance fluorouracil and leucovorin. She developed further peritoneal progression including a metastasis to a periumbilical area that extended through the skin obliterating the umbilicus. mFOLFOX was reinstituted but after 6 cycles of treatment there was further tumor growth. To identify avenues for potential benefit from targeted therapy, the patient's tumor tissue obtained from the colectomy specimen was submitted for comprehensive genomic profiling by clinical NGS utilizing the FoundationOne platform.

Comprehensive Genomic Profiling

Clinical NGS-based comprehensive genomic profiling was performed using DNA extracted from formalin-fixed paraffin-embedded sections cut at 10 μπι thickness in a CLIA-certified laboratory (Foundation Medicine, Cambridge, MA). The sample forwarded for DNA extraction contained a minimum of 20% DNA derived from tumor cells. DNA sequencing was performed for the entire coding region of 315 cancer-related genes plus introns from 28 genes frequently rearranged in cancer on an indexed, adaptor- ligated, hybridization-captured library and fully sequenced using 49 bp reads (Illumina HiSeq 2500, Hayward, CA) to a median exon coverage of 555X for this specific case. The bait set used in sequencing included key genes involved in colorectal carcinogenesis, such as KRAS, BRAF, PIK3CA, PTEN, CTNNB 1, APC, and MMR MLH1, PMS2, MSH2 and MSH6. Sequence reads were mapped to the reference human genome (hgl9) and analyzed for genomic alterations including base substitutions, small insertions and deletions (indels), copy number alterations (amplifications and homozygous deletions), and select gene fusions/rearrangements as previously described (Frampton GM, Fichtenholtz A, Otto GA, et al. Development and validation of a clinical cancer genomic profiling test based on massively parallel DNA sequencing. Nat Biotechnol 2013;31 : 1023-1031.).

Immunohistochemistry

Immunohistochemistry was performed on 4-μπι formalin-fixed paraffin-embedded tissues sections using a BenchMark Ultra instrument, version 4 (Ventana Medical Systems, Tucson, AZ). The primary rabbit monoclonal anti-ALK antibody (clone D5F3; Cell Signaling Technology, Beverly, MA) was used at a 1 :50 dilution. Fluorescence in situ hybridization (FISH)

Fluorescence in situ hybridization (FISH) was performed on 4-μιη paraffin sections using the Vysis LSI ALK Dual Color Break Apart rearrangement probe (Abbott Molecular, Abbott Park, IL).

RESULTS

Comprehensive genomic profiling, immunihistochemistry. and fish

Comprehensive genomic profiling of the tumor sample identified 14 genomic alterations: a STR -ALK fusion (Fig. 2, panel C) and mutations in KRAS R164Q, STK11, TP53, AEID1A, BCOR, CDH2, MLL2, MLL3, PAX5, and RAD50 genes. Although no BRAF mutations were identified, genomic alterations were not detected in genes encoding the mismatch repair proteins MLH1, PMS2, MLH2, and MLH6, consistent with a sporadic microsatellite instable (MSI) CRC. The KRAS R164Q mutation identified in this case is outside of the "hotspot" codons and is phenotypically equivalent to wild type KRAS.

A STRN-ALK fusion involved the intrachromosomal translocation of exons 1-3 of STRN to exons 20-29 of ALK (Fig. 2, panel D). Both fusion partners are located on the short arm of chromosome 2 (2p22.2 and 2p23, separated by ~ 7.69 Mb), indicating that the fusion is a result of intrachromosomal paracentric rearrangement. The predicted fusion protein does not have a calmodulin-binding domain and the WD-repeats, but retains the N-terminal caveolin-binding and the coiled-coil binding domains of STRN, fused to the intracellular juxtamembrane region of ALK, containing the kinase domain (Fig. 2, panel D). The coiled-coil STRN domain may act as a dimerization motif that could constitutively activate ALK tyrosine kinase.

Immunohistochemical analysis demonstrated diffuse intracytoplasmic immunoreactivity with the anti-ALK antibody (Fig. 3, panel E). This staining pattern confirmed the loss of ALK extracellular and intramembrane domains as detected by sequencing, and thus the positioning of the fusion protein in the intracellular compartment.

FISH analysis confirmed the presence of ALK gene rearrangement in 74% of the tumor cells (Fig 3, panel F).

Clinical response to ceritinib

Based on the STRN-ALK fusion, the patient received ceritinib, an orally available ALK inhibitor, 750 mg/day, on the Novartis Signature Trial. Periumbilical skin metastasis significantly decreased within 7 weeks of treatment (Fig.4 B). After an additional 2 weeks of treatment there was marked decrease in periumbilical thickening (Fig.4 C). At 6 month follow-up the mass protruding through the skin can no longer be seen (Fig.4 D) and there has been resolution of all contrast enhancing tumor by CT scan (Fig.4 E-F). The patient currently remains on active treatment.

Conclusions:

A patient was reported with progressive metastatic colon cancer with a novel STRN-ALK gene fusion identified by comprehensive genomic profiling. The patient had a dramatic response to the tyrosine kinase ALK inhibitor ceritinib and remains on treatment after 6 months of therapy. This is the first case report of a transforming ALK fusion as a therapeutic target in colorectal cancer and illustrates the potential for personalized molecular based therapy for the treatment of advanced malignancies.

Claims

1. An ALK inhibitor for use in treating colorectal cancer characterized by the presence of a STRN-ALK gene fusion or a STR -ALK fusion polypeptide.

2. The ALK inhibitor for use in treating colorectal cancer according to claim 1, wherein the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide has been detected in a biological sample from the cancer obtained from a patient having said cancer.

3. The ALK inhibitor for use in treating a patient having colorectal cancer according to claim 2 characterized in that:

(a) the patient is selected for the treatment with an ALK inhibitor on the basis of the patient having STRN-ALK gene fusion or the STRN-ALK fusion polypeptide; and

(b) thereafter, a therapeutically effective amount of an ALK inhibitor is

administered to the patient.

4. The ALK inhibitor for use in treating a patient having colorectal cancer according to claim 2 or claim 3, characterized in that:

(b) a therapeutically effective amount of an ALK inhibitor is selectively

administered to the patient on the basis of the biological sample from the patient having STRN-ALK gene fusion or the STRN-ALK fusion polypeptide.

5. Use of an ALK inhibitor for the preparation of a medicament for the treatment of colorectal cancer, wherein the colorectal cancer is characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide.

6. A method of selectively treating a patient having colorectal cancer with an ALK inhibitor, wherein the method comprises the steps of:

(a) selecting the patient for the treatment with an ALK inhibitor on the basis of the patient having the colorectal cancer characterized by the presence of a STRN- ALK gene fusion or a STRN-ALK fusion polypeptide; and

7. A method of selectively treating a patient having colorectal cancer, comprising either:

(a) selectively administering a therapeutically effective amount of an ALK inhibitor to the patient on the basis of the patient having the colorectal cancer characterized by the presence of a STRN-ALK gene fusion or a STR -ALK fusion polypeptide; or

(b) selectively administering a therapeutically effective amount of a drug other than an ALK inhibitor to the patient on the basis of the patient not having the colorectal cancer characterized by the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide.

8. A method of selectively treating a patient having colorectal cancer with an ALK inhibitor, comprising:

(c) thereafter, administering a therapeutically effective amount of an ALK

inhibitor to the patient.

9. A method of selectively treating a patient having colorectal cancer, comprising:

(b) thereafter, selectively administering to the patient either:

(ii) a therapeutically effective amount of a drug other than an ALK inhibitor on the basis of the biological sample obtained from the patient not having the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide.

10. A method of predicting the likelihood that a patient having a colorectal cancer will respond to treatment with an ALK inhibitor, comprising assaying a biological sample obtained from the patient for the presence or absence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide, wherein:

(a) the presence of the STRN-ALK gene fusion or the STRN-ALK fusion

polypeptide is indicative of an increased likelihood that the patient will respond to treatment with an ALK inhibitor; and (b) the absence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide is indicative of a decreased likelihood that the patient will respond to treatment with an ALK inhibitor.

11. A method of characterizing a human colorectal cancer, comprising

(a) Obtaining a biological sample from said human colorectal cancer; and

(b) Detecting the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide in said sample, thereby characterizing said colorectal cancer based on the presence or absence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide.

A method for detecting the presence of the STRN-ALK gene fusion or the STRN- ALK fusion polypeptide in a biological sample from a human colorectal cancer, comprising detecting the presence of the STRN-ALK gene fusion or the STRN- ALK fusion polypeptide in said sample, thereby detecting whether the STRN- ALK gene fusion or the STRN-ALK fusion polypeptide is present in said biological sample.

The method of characterizing a human colorectal cancer according to claim 11 or method for detecting according to claim 12, wherein the presence of the STRN- ALK gene fusion or the STRN-ALK fusion polypeptide is detected in the sample obtained from a patient having said colorectal cancer.

A kit for detecting the presence of a STRN-ALK gene fusion or a STRN-ALK fusion polypeptide in a biological sample from a human colorectal cancer, comprising at least one probe capable of detecting the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide and instructions for using the probe to assay the biological sample for the presence of the STRN-ALK gene fusion or the STRN-ALK fusion polypeptide.

A kit for use in predicting the likelihood that a patient having colorectal cancer will respond to the treatment with an ALK inhibitor comprising:

16. A kit for use in treating a patient having colorectal cancer comprising:

(a) a therapeutically effective amount of an ALK inhibitor;

(d) instructions for administering the ALK inhibitor to the patient if the STRN- ALK gene fusion or the STRN-ALK fusion polypeptide is present in the biological sample obtained from the patient ; and

(e) optionally, means for administering the ALK inhibitor to the patient.

17. The kit according to any one of claims 14 to 16, wherein the probe is an oligonucleotide that specifically hybridizes to a region of a nucleic acid coding for the STRN-ALK gene fusion, or an oligonucleotide that specifically hybridizes to a region of a nucleic acid coding for an equivalent genetic marker of the STRN- ALK gene fusion, or the probe is an antibody that binds to the STRN-ALK fusion polypeptide.

18. The ALK inhibitor for use according to any one of claims 1 to 4 or the method according to any one of claims 6 to 10, the method of characterizing a human colorectal cancer according to claim 11 or claim 13, the method for detecting according to claim 12 or claim 13, or a kit according to anyone of claims 14 to 17, wherein the assaying or detecting for STRN-ALK gene fusion or the STRN-ALK fusion polypeptide comprises a technique selected from the group consisting of Next Generation Sequencing (NGS), Northern blot analysis, polymerase chain reaction (PCR), reverse transcription-polymerase chain reaction (RT-PCR), TaqMan-based assays, direct sequencing, dynamic allele-specific hybridization, high-density oligonucleotide SNP arrays, restriction fragment length polymorphism (RFLP) assays, primer extension assays, oligonucleotide ligase assays, Southern Blot, immunoassays, immunohistochemistry, ELISA, fluorescence in situ hybridization analysis (FISH), kinase activity assays, flow cytometry, Western blot, HPLC, and mass spectrometry.

19. The ALK inhibitor for use according to any one of claims 1 to 4 or the method according to any one of claims 6 to 13, or a kit according to anyone of claims 14 to 17, wherein the step of assaying or detecting for STRN-ALK gene fusion comprises PCR, RT-PCR or fluorescence in situ hybridization analysis (FISH).

20. The ALK inhibitor for use according to any one of claims 1 to 4 or the method according to any one of claims 6 to 13, or a kit according to anyone of claims 14 to 17, wherein the step of assaying or detecting for the STRN-ALK fusion polypeptide comprises immunohistochemistry.

21. The ALK inhibitor for use according to any one of claims 1-4, 18 to 20, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 5 to 10, or 18 to 20, or a kit according to anyone of claims 15 to 20, wherein said inhibitor is selected from the group consisting of

CrcroJ.Wb

and 5-chloro-N2-(2-isopropoxy-5-methyl-4-(piperidin-4-yl)phenyl)-N4-[2- (propane-2-sulfonyl)-phenyl]-pyrimidine-2,4-diamine, or a pharmaceutically acceptable salt thereof.

22. The ALK inhibitor for use according to any one of claims 1-4, 18 to 21, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 5 to 10, or 18 to 21, or a kit according to anyone of claims 15 to 21, wherein said inhibitor is 5 -chloro-N2-(2-isopropoxy-5 -methyl -4-(piperidin-4- yl)phenyl)-N4-[2-(propane-2-sulfonyl)-phenyl]-pyrimidine-2,4-diamine, or a pharmaceutically acceptable salt thereof.

23. The ALK inhibitor for use according to any one of claims 1-4, 18 to 22, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 6 to 13, or 18 to 22, or a kit according to anyone of claims 14 to 22, wherein said colorectal cancer is metastatic.

24. The ALK inhibitor for use according to any one of claims 1-4, 18 to 23, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 6 to 13, or 18 to 23, or a kit according to anyone of claims 14 to 23, wherein the ALK inhibitor is administered for at least 6 months.

25. The ALK inhibitor for use according to any one of claims 1-4, 18 to 23, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 6 to 13, or 18 to 24, or a kit according to anyone of claims 14 to 23, wherein the ALK inhibitor leads to a progression free survival of at least 6 months.

26. The ALK inhibitor for use according to any one of claims 1-4, 18 to 25, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 6 to 13, or 18 to 25, or a kit according to anyone of claims 14 to 25, where the ALK inhibitor is administered as a second line treatment.

27. The ALK inhibitor for use according to any one of claims 1-4, 14 to 21, or the use of the ALK inhibitor according to claim 5, or the method according to any one of claims 6 to 13, or 18 to 25, or a kit according to anyone of claims 14 to 25, where the ALK inhibitor is administered as a first line treatment.