METHODS FOR THE IDENTIFICATION, EVALUATION AND TREATMENT OF PATIENTS HAVING MULTIPLE MYELOMA

METHODES D'IDENTIFICATION, D'EVALUATION ET DE TRAITEMENT DES PATIENTS ATTEINTS DE MYELOMES MULTIPLES

English Abstract

The present disclosure relates to methods for the treatment of multiple
myeloma in
patients who have one or more cytogenetic alterations. In particular, the
disclosure provides
methods for treatment of multiple myeloma in patients who have one or more
cytogenetic
alternations by administering to a patient a treatment regimen comprising a
proteasome
inhibitor or a pharmaceutically acceptable salt, stereoisomeric or tautomeric
form thereof if
said patient is identified as a likely responder to the treatment regimen by
assessing the
presence of said cytogenetic alterations.

Claims

CLAIMS:
1. A method of treating a patient having multiple myeloma, comprising:
(i) determining if the patient has a cytogenetic alteration at chromosome 17,
and
(ii) if the patient has a cytogenetic alteration at chromosome 17, then
administering to
a patient in need thereof a treatment regimen comprising a compound of formula
(Ia):
Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
2. The method of claim 1, wherein the cytogenetic alteration at chromosome
17 is del
(17).
3. The method of claim 2, wherein the deletion at chromosome 17 is del
(17p13).
4. The method of any one of claims 1-3, wherein the patient is determined
to also have at
least one other cytogenetic alteration wherein the cytogenetic alteration is
t(4:14).
5. The method of any one of claims 1-3, wherein the patient is determined
to also have at
least one other cytogenetic alteration wherein the cytogenetic alteration is
t(14:16).
6. The method of any one of claims 1-3, wherein the patient is determined
to also have at
least two other cytogenetic alterations wherein the cytogenetic alterations
are t(4:14) and
t(14:16).
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7. The method of any one of claims 1-3, wherein the determining if a
patient has a
cytogenetic alteration at chromosome 17 comprises the steps of:
(i) providing a bone marrow aspirate sample from the patient;
(ii) isolating CD138 positive plasma cells from the sample; and
(iii) performing FISH analysis on the CD138 enriched positive plasma cells.
8. The method of any one of claims 1-7, wherein the compound of formula
(Ia) is
characterized as the compound of formula (IIIa):
Image
or a pharmaceutically acceptable salt thereof.
9. The method of claim 8, wherein the compound of formula (IIIa) is in a
solid dosage
form and the solid dosage form is a capsule.
10. The method of claim 9, wherein the capsule comprises a mixture of the
compound of
formula (IIIa) or a pharmaceutically acceptable salt thereof, microcrystalline
cellulose, talc,
and magnesium stearate.
11. The method of any one of claims 1-10 wherein the compound of formula
(Ia) is
administered on each of days 1, 8 and 15 of a 28 day cycle.
12. The method of any one of claims 1-11, wherein the treatment regimen
further
comprises lenalidomide and dexamethasone.
13. A method of treating a patient having a multiple myeloma, comprising:
i) selecting a patient based upon the patient having a cytogenetic
alteration at
chromosome 17; and
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ii) administering to the patient a treatment regimen comprising a
compound of
formula (Ia):
Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
14. The method of claim 13, wherein the cytogenetic alteration at
chromosome 17 is del
(17).
15. The method of claim 13, wherein the deletion at chromosome 17 is del
(17p13).
16. The method of any one of claims 13-15, wherein the patient is
determined to also have
at least one other cytogenetic alteration wherein the cytogenetic alteration
is t(4:14).
17. The method of any one of claims 13-15, wherein the patient is
determined to also have
at least one other cytogenetic alteration wherein the cytogenetic alteration
is t(14:16).
18. The method of any one of claims 13-15, wherein the patient is
determined to also have
at least two other cytogenetic alterations wherein the cytogenetic alterations
are t(4:14) and
t(14:16).
19. The method of any one of claims 13-15, wherein the determining if a
patient has a
cytogenetic alteration at chromosome 17 comprises the steps of:
(i) providing a bone marrow aspirate sample from the patient;
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(ii) isolating CD138 positive plasma cells from the sample; and
(iii) performing FISH analysis on the CD138 enriched positive plasma cells.
20. The method of any one of claims 13-19, wherein the compound of formula
(Ia) is
characterized as the compound of formula (IIIa):
Image
or a pharmaceutically acceptable salt thereof
21. The method of claim 20, wherein the compound of formula (IIIa) is in a
solid dosage
form and the solid dosage form is a capsule.
22. The method of claim 21, wherein the capsule comprises a mixture of the
compound of
formula (IIIa) or a pharmaceutically acceptable salt thereof, microcrystalline
cellulose, talc,
and magnesium stearate.
23. The method of any one of claims 13-22, wherein the compound of formula
(Ia) is
administered on each of days 1, 8 and 15 of a 28 day cycle.
24. The method of any one of claims 13-23, wherein the treatment regimen
further
comprises lenalidomide and dexamethasone.
25. A method of evaluating a patient having multiple myeloma for
responsiveness to a
treatment regimen comprising a compound of formula (Ia):
Image
- 41 -

or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O;
comprising the steps:
i) determining whether a patient has a cytogenetic alteration at chromosome
17;
ii) recording the presence or absence of a cytogenetic alteration at
chromosome
17, and
iii) determining, recommending or selecting an appropriate treatment
regimen
based upon the presence or absence of a cytogenetic alteration at chromosome
17.
26. The method of claim 25, wherein step iii) comprises determining whether
to begin or
continue the treatment regimen comprising the compound of formula (Ia) or a
pharmaceutically acceptable salt, stereoisomeric or tautomeric form thereof, ,
based upon the
presence of a cytogenetic alteration at chromosome 17.
27. An article of manufacture comprising:
i) a compound of formula (Ia):
Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof,
wherein Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form
a cyclic
boronic ester having 2-20 carbon atoms, and optionally one or more heteroatoms
selected
from N, S, or O; and
ii) instructions for determining the appropriateness of use of said
composition by
determining whether a patient has a cytogenetic alteration at chromosome 17.
28. An article of manufacture comprising:
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i) reagents for making a determination whether a patient has a cytogenetic
alteration at chromosome 17, and
ii) instructions for determining the appropriateness of use of a
pharmaceutical
composition comprising a compound of formula (Ia):
Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof,
wherein Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form
a
cyclic boronic ester having 2-20 carbon atoms, and optionally one or more
heteroatoms selected from N, S, or O on the basis of said determination.
29. An article of manufacture comprising:
i) a pharmaceutical composition comprising a compound of formula (Ia):
Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein Z1 and Z2 are each independently hydroxyl; or Z1 and Z2
together form a cyclic boronic ester haying 2-20 carbon atoms, and
optionally one or more heteroatoms selected from N, S, or O;
ii) reagents for making a determination whether a patient has a cytogenetic

alteration at chromosome 17; and
iii) instructions for determining the appropriateness of use of a
pharmaceutical
composition comprising a compound of formula (Ia) or a pharmaceutically
acceptable salt, stereoisomeric or tautomeric form on the basis of said
determination.
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30. Use of a compound of formula I for treating a patient having multiple
myeloma,
wherein the use comprises:
determining if the patient has a cytogenetic alteration at chromosome 17, and
(ii) if the patient has a cytogenetic alteration at chromosome 17,
selecting the
patient for administration of a treatment regimen comprising a compound of
formula (la):
Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
31. The use of claim 30, wherein the cytogenetic alteration at chromosome
17 is del (17).
32. The use of claim 31, wherein the deletion at chromosome 17 is del
(17p13).
33. The use of any one of claims 30-32, wherein the patient is determined
to also have at
least one other cytogenetic alteration wherein the cytogenetic alteration is
t(4:14).
34. The use of any one of claims 30-32, wherein the patient is determined
to also have at
least one other cytogenetic alteration wherein the cytogenetic alteration is
t(14:16).
35. The use of any one of claims 30-32, wherein the patient is determined
to also have at
least two other cytogenetic alterations wherein the cytogenetic alterations
are t(4:14) and
t(14:16).
36. The use of any one of claims 30-32, wherein the determining if a
patient has a
cytogenetic alteration at chromosome 17 comprises the steps of:
providing a bone marrow aspirate sample from the patient;
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(ii) isolating CD138 positive plasma cells from the sample; and
(iii) performing FISH analysis on the CD138 enriched positive plasma cells.
37. The use of any one of claims 30-36, wherein the compound of formula
(Ia) is
characterized as the compound of formula (IIIa):
Image
or a pharmaceutically acceptable salt thereof.
38. The use of claim 37, wherein the compound of formula (IIIa) is in a
solid dosage form
and the solid dosage form is a capsule.
39. The use of claim 38, wherein the capsule comprises a mixture of the
compound of
formula (IIIa) or a pharmaceutically acceptable salt thereof, microcrystalline
cellulose, talc,
and magnesium stearate.
40. The use of any one of claims 30-39 wherein the compound of formula (Ia)
is for
administration on each of days 1, 8 and 15 of a 28 day cycle.
41. The use of any one of claims 30-40, wherein the treatment regimen
further comprises
lenalidomide and dexamethasone.
42. Use of a compound of formula (Ia) for treating a patient having
multiple myeloma,
comprising:
i) selecting a patient, based upon the patient having a cytogenetic
alteration at
chromosome 17, for administration of a treatment regimen comprising a compound
of
formula (Ia):
- 45 -

Image
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
43. The use of claim 42, wherein the cytogenetic alteration at chromosome
17 is del (17).
44. The use of claim 42, wherein the deletion at chromosome 17 is del
(17p13).
45. The use of any one of claims 42-44, wherein the patient is determined
to also have at
least one other cytogenetic alteration wherein the cytogenetic alteration is
t(4:14).
46. The use of any one of claims 42-44, wherein the patient is determined
to also have at
least one other cytogenetic alteration wherein the cytogenetic alteration is
t(14:16).
47. The use of any one of claims 42-44, wherein the patient is determined
to also have at
least two other cytogenetic alterations wherein the cytogenetic alterations
are t(4:14) and
t(14:16).
48. The use of any one of claims 42-44, wherein the determining if a
patient has a
cytogenetic alteration at chromosome 17 comprises the steps of:
providing a bone marrow aspirate sample from the patient;
(ii) isolating CD138 positive plasma cells from the sample; and
(iii) performing FISH analysis on the CD138 enriched positive plasma cells.
49. The use of any one of claims 42-48, wherein the compound of formula
(Ia) is
characterized as the compound of formula (IIIa):
- 46 -

Image
or a pharmaceutically acceptable salt thereof.
50. The use of claim 49, wherein the compound of formula (IIIa) is in a
solid dosage form
and the solid dosage form is a capsule.
51. The use of claim 50, wherein the capsule comprises a mixture of the
compound of
formula (IIIa) or a pharmaceutically acceptable salt thereof, microcrystalline
cellulose, talc,
and magnesium stearate.
52. The use of any one of claims 42-51, wherein the compound of formula
(Ia) is for
administeration on each of days 1, 8 and 15 of a 28 day cycle.
53. The use of any one of claims 42-52, wherein the treatment regimen
further comprises
lenalidomide and dexamethasone.
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Description

CA 02915105 2015-12-10
METHODS FOR THE IDENTIFICATION, EVALUATION AND TREATMENT OF
PATIENTS HAVING MULTIPLE MYELOMA
FIELD
[001] The present disclosure relates to methods for the treatment of
multiple myeloma in
patients who have one or more cytogenetic alterations. In particular, the
disclosure provides
methods for treatment of multiple myeloma in patients who have one or more
cytogenetic
alterations by administering to a patient a treatment regimen comprising a
proteasome
inhibitor or a pharmaceutically acceptable salt, stereoisomeric or tautomeric
form thereof, if
said patient is identified as a likely responder to the treatment regimen by
assessing the
presence of said cytogenetic alterations.
BACKGROUND
[002] Multiple myeloma, a B-cell tumor of malignant plasma cells within the
bone
marrow, remains incurable despite advances in novel therapies with proteasome
inhibitors
(PIs), immunomodulating drugs (IMiD), and stem cell transplant (SCT) therapy.
Multiple
myeloma is characterized by the accumulation of plasma cells in the bone
marrow (and other
organs) and can result in bone marrow failure, bone destruction,
hypercalcemia, and renal
failure. It constitutes approximately 1% of all reported neoplasms and
approximately 13% of
hematologic cancers worldwide. In the Americas, Canada, and Western European
countries,
approximately five to seven new cases of multiple myeloma are diagnosed per
100,000 people
each year. Palumbo and Anderson, N Engl J Med 2011;364(11):1046-60; Landgren
and
Weiss, Leukemia 2009;23(10):1691-7; Harousseau, et al., Annals of Oncology
2008;19 Suppl
2:ii55-7. Although less common in Asian countries, incidences of multiple
myeloma have
increased almost 4-fold in the past 25 years and are characterized by younger
age of onset,
more invasive disease, and a less favorable prognosis (Huang, et al., Cancer
2007;110(4):896-
905; Qiu, et al., Clinical Epidemiological Study on Multiple Myeloma in China
(ASH Annual
Meeting Abstracts) 2008;112(11):abstr 2723).
[003] Multiple myeloma is sensitive to many cytotoxic drugs including
alkylating
agents, anthracyclines, and corticosteroids for both initial treatment and
relapsed disease.
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CA 02915105 2015-12-10
Over the past decade, significant achievements have been made in expanding
treatment
options for multiple myeloma with novel therapies such as thalidomide,
bortezomib, and
lenalidomide. These regimens have extended progression-free survival (PFS)
and/or time-to-
progression (TTP) (Palumbo, et al., Leukemia 2008; 22(2):414-23; Mateos, et
al., Journal of
Clinical Oncology 2010; 28(13):2259-66; Gay, et al., Haematologica 2010;
94:0507;
Richardson, et al., Hematology 2007:317-23; Dimopoulos, et al., Leukemia 2009;

23(11):2147-52). The introduction of novel therapies and the increased use of
high-dose
therapy (HDT) significantly improved overall survival in patients with newly
diagnosed
multiple myeloma (NDMM) who were eligible for autologous stem cell transplant
(ASCT)
(Kumar, et al., Blood 2008; 111(5):2516-20; Brenner, et al., Blood 2008;
111(5):2521-6;
Libby, et al., Declining myeloma mortality rates in the United States
following introduction of
novel therapies In: International Myeloma Workshop Paris, France; 2011).
[004] Despite more therapeutic options, multiple myeloma remains incurable,
and patients
with early stage cancer remain at risk for relapse after their initial
therapy. When patients
relapse after their initial therapy, they demonstrate variable responses to
subsequent treatments
with decreasing likelihood and duration of response (DOR). Patients become
refractory to
approved therapies and ultimately are left with no alternative treatment
options. Importantly,
the survival of a subgroup of patients with certain cytogenetic abnormalities
or alterations
(collectively referred to as high-risk multiple myeloma) has remained poor
despite aggressive
therapy incorporating almost every available drug and treatment modality.
Therefore, there
remains a need for new and better drugs and regimens or strategies to overcome
high-risk
prognostic factors and improve response rates and survival rates in this
patient population.
DESCRIPTION
[005] Unless otherwise defined, all technical and scientific terms used
herein have the
same meaning as commonly understood by one of ordinary skill in the art to
which this
disclosure belongs.
[006] Terms used herein shall be accorded the following defined meanings,
unless
otherwise indicated.
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CA 02915105 2015-12-10
[007] "Treatment" shall mean the use of a therapy to prevent or inhibit
further tumor
growth, as well as to cause shrinkage of a tumor or tumor burden, and to
provide longer
survival times. Treatment is also intended to include prevention of metastasis
of tumor. A
tumor is "inhibited" or -treated" if at least one symptom (as determined by
responsiveness/non-responsiveness, time to progression, or indicators known in
the art and
described herein) of the cancer or tumor is alleviated, terminated, slowed,
minimized, or
prevented.
[008] "Treatment regimen" as used herein, refers to treatment with a
molecule alone, or
in combination with another molecule. A treatment regimen also refers to dose
amount, the
frequency of dosing and the number of times a molecule, or combination of
molecules, is
administered.
[009] As used herein, a "favorable" outcome or prognosis refers to long
term survival,
long time-to-progression (TTP), and/or good response. Conversely, an
"unfavorable"
outcome or prognosis refers to short term survival, short time-to-progression
(TTP) and/or
poor response.
[0010] As used herein, "long time-to-progression, "long TTP" and "short
time-to-
progression," "short TTP" refer to the amount of time until when the stable
disease brought
by treatment converts into an active disease. On occasion, a treatment results
in stable disease
which is neither a good nor a poor response, e.g., MR, the disease merely does
not get worse,
e.g., become a progressive disease, for a period of time. This period of time
can be at least 4-
8 weeks, at least 3-6 months or more than 6 months.
[0011] The term "survival" refers to the patient remaining alive, and
includes progression-
free survival (PFS) and overall survival (OS). Survival can be estimated by
the Kaplan-Meier
method, and any differences in survival are computed using the stratified log-
rank test.
[0012] The term "progression-free survival (PFS)" refers to the time from
treatment (or
randomization) to first disease progression or death. For example it is the
time that the patient
remains alive, without return of the cancer (e.g., for a defined period of
time such as about
one month, two months, three months, three and a half months, four months,
five months, six
months, seven months, eight months, nine months, about one year, about two
years, about
three years, about five years, about 10 years, about 15 years, about 20 years,
about 25 years,
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CA 02915105 2015-12-10
etc.) from initiation of treatment or from initial diagnosis. Progression free
survival can be
measured in multiple myeloma by using International Myeloma Working Group
(IMWG)
criteria.
[0013] The term "overall survival" refers to the patient remaining alive
for a defined
period of time (such as about one year, about two years, about three years,
about four years,
about five years, about 10 years, about 15 years, about 20 years, about 25
years, etc.) from
initiation of treatment or from initial diagnosis.
[0014] The term "proteasome-mediated disorder" refers to any disorder,
disease or
condition which is caused or characterized by an increase in proteasome
expression or
activity, or which requires proteasome activity. The term "proteasome-mediated
disorder"
also includes any disorder, disease or condition in which inhibition of
proteasome activity is
beneficial. Proteasome inhibitors are drugs that block the action of
proteasomes, cellular
complexes that break down proteins, like the p53 protein. Proteasome
inhibitors are being
studied in the treatment of cancer, especially multiple myeloma. Examples of
proteasome
inhibitors are bortezomib, carfilzomib, ixazomib, disulfiram, epigallocatechin-
3-gallate,
salinosporamid A, 0NX0912, CEP-18770, and epoxomicin.
[0015] The term "about" is used herein to mean approximately, in the region
of, roughly,
or around. When the term "about" is used in conjunction with a numerical
range, it modifies
that range by extending the boundaries above and below the numerical values
set forth. In
general, the term "about" is used herein to modify a numerical value above and
below the
stated value by a variance of 10%.
[0016] The term "comprises" refers to "includes, but is not limited to."
[0017] The term "pharmaceutically acceptable carrier" is used herein to
refer to a material
that is compatible with a recipient subject, preferably a mammal, more
preferably a human,
and is suitable for delivering an active agent to the target site without
terminating the activity
of the agent. The toxicity or adverse effects, if any, associated with the
carrier preferably are
commensurate with a reasonable risk/benefit ratio for the intended use of the
active agent.
[0018] The term "orally" refers to administering a composition that is
intended to be
, ingested. Examples of oral forms include, but are not limited to, tablets,
pills, capsules,
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CA 02915105 2015-12-10
powders, granules, solutions or suspensions, and drops. Such forms may be
swallowed whole
or may be in chewable form.
[0019] The term "patient", as used herein, means an animal, preferably a
mammal, more
preferably a human.
[0020] A -cytogenetic alteration" is a chromosomal change such as a
deletion,
duplication, or translocations in a chromosome.
[0021] A "kit" is any article of manufacture (e.g., a package or container)
comprising at
least one reagent, e.g. a probe, for specifically detecting one or more
cytogenetic alterations of
the disclosure. The article of manufacture may be promoted, distributed, sold
or offered for
sale as a unit for performing, e.g., in vitro, the methods of the present
disclosure, e.g., on a
sample having been obtained from a patient. The reagents included in such a
kit can comprise
probes/primers and/or antibodies for use in detecting one or more cytogenetic
alterations. In
addition, a kit of the present disclosure can contain instructions which
describe a suitable
detection assay. Such a kit can be conveniently used, e.g., in a clinical or a
contract testing
setting, to generate information to be recorded, stored, transmitted or
received to allow for
diagnosis, evaluation or treatment of patients exhibiting symptoms of multiple
myeloma.
[0022] As used herein, the term "evaluating a patient" refers to the act of
reviewing or
analyzing a patient's cytogenetic alteration. The evaluation can further
include one or more
of: obtaining a sample from a patient (e.g. a sample from a bodily fluid (e.g.
a blood sample,
a serum sample, a urine sample, a synovial fluid sample, a tear sample, a
saliva sample) or a
tissue sample (e.g., a skin sample or a tissue sample obtained from a biopsy)
or analyzing a
sample in vivo; assaying the sample or requesting an assay using the sample to
obtain
genomic information regarding the patient's cytogenetic alteration; reviewing
the patient's
information using the assay results performed with the sample and/or a
patient's medical
records. The patient's information (e.g. genomic information or value on the
patient's
cytogenetic alteratioin) can then optionally be compared to a reference
standard, e.g., publicly
available information (i.e. against a reference population), to make an
informed decision
regarding treatment options for that patient.
[0023] The terms "boronate ester" and "boronic ester" are used
interchangeably and refer
to a chemical compound containing a -B(Z1)(Z2) moiety, wherein Z1 and Z2
together form a
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CA 02915105 2015-12-10
cyclic boronic ester having 2-20 carbon atoms, and optionally one or more
heteroatoms
selected from N, S, or O.\
[0024] In some embodiments, the present disclosure provides a method of
treating a
patient having multiple myeloma, comprising:
(i) determining if the patient has a cytogenetic alteration at chromosome
17, and
(ii) if the patient has a cytogenetic alteration at chromosome 17, then
administering
to a patient in need thereof a treatment regimen comprising a compound of
formula (/):
0 Z1
H i
0N =rNi Bz2
H
0
Y (1)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein
ring A is
F
1110 121 CI tio 1%,
40 F 0 14_ 40
r lk
F \ 0
B
9 9 9 9 7
CI
F io 11/4.
F
40 \ 40
, CI CI CI
9 110 40
F CI 9 9 9 F / F 9
so \
c,
40 \ 40 \ = 40 40 µ
c, Cl,F F CI F CI CI CI
, , , , , or
c, io
Cl ;and
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
[0025] In some embodiments, the compound of formula (/) is characterized by
formula
(Ia):
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CA 02915105 2015-12-10
ci 0
N NIE3=z2
0
CI (Ia)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroato.ms selected
from N, S, or O.
[0026] In some embodiments, the present disclosure provides a method of
treating a
patient having multiple myeloma, comprising:
(i) determining if the patient has a cytogenetic alteration at chromosome 17,
and
(ii) if the patient has a cytogenetic alteration at chromosome 17, then
administering to
a patient in need thereof a treatment regimen comprising a compound of formula
(la):
CI
N'NB
= r z2
0
Cl (Ia)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
[0027] In some embodiments, the compound of formula (I) is characterized by
formula
(//):
0
0 =
9A,(1.1/7
R1
N.rNE3.0 R2
0
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
ring A is defined above; R1 and R2 independently is -(CH2)p-CO2H; wherein one
of carboxylic
acids optionally forms a further bond with the boron atom; n is 0 or 1; and p
is 0 or 1.
[0028] In some embodiments, the compound of formula (/) is characterized by
formula
(///):
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CA 02915105 2015-12-10
o
0 [1:3:310 C 02 H
0
CO2H
(HI)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein ring
A is defined above.
[0029] In some embodiments, the compound of formula (/) is characterized by
formula
(IHa):
0
CI 0 o-(= co2H
0
CO2H
CI (IHa)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof.
[0030] In some embodiments, the compound of formula (I) is characterized by
formula
(/V):
CI 0 OH
(10
Nr \/LOH
0
CI (/V)
or an ester or a pharmaceutically acceptable salt thereof
[0031] The compound of formula (/V), also known as ixazomib, is a peptide
boronic acid
developed by Millennium Pharmaceuticals, Inc. Ixazomib is a biologically
active molecule
that potently, reversibly, and selectively inhibits the proteasome. The
compound of formula
(IHa) is a citrate ester of ixazomib, referred to as ixazomib citrate herein.
Ixazomib citrate
rapidly hydrolyzes to ixazomib upon contact with either plasma or aqueous
solutions.
[0032] In some embodiments, the compound of formulas (/), (Ia), (II),
(III). (IHa) or
(/V) are administered orally. In some embodiments, the compound of formula
(/), (Ia),
(III). (IHa) or (/V) is administered in a solid dosage form. In some
embodiments, the solid
dosage form is a capsule. In some embodiment, the capsule comprises a mixture
of the
compound of formula (IIIa) or a pharmaceutically acceptable salt thereof,
micryocrystalliine
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CA 02915105 2015-12-10
cellulose, talc and magnesium stearate. In some embodiments the capsule
comprises a
mixture of the compound of formula (lila), microcrystalline cellulose, talc
and magnesium
stearate.
[0033] Synthetic methods for the preparation of compounds and
pharmaceutical
compositions of formulas (/), (Ia), (II), (III), (IHa) and (IV) are, for
example, described in
US Patent 7,442,830, US Patent 7,687,662, US Patent 8,003,819, US Patent
8,530,694, and
International Patent Publication WO 2009/154737, which are hereby incorporated
by
reference specifically and in their entirety.
[0034] In some embodiments, the compound of formula (/) is administered on
each of
days 1, 8 and 15 of a 28 day cycle. In some embodiments, the compound of
formula (Ia) is
administered on each of days 1, 8 and 15 of a 28 day cycle. In some
embodiments, the
compound of formula (IHa) is administered on each of days 1, 8 and 15 of a 28
day cycle. In
some embodiments, the compound of formula (/V) is administered on each of days
1, 8 and
15 of a 28 day cycle.
[0035] In some embodiments, the amount of the compound of formula (IV) that
is
administered is 4 mg, 3 mg or 2.3 mg. In some embodiments, the amount of the
compound of
formula (/V) that is administered is 4 mg. In some embodiments, the amount of
the
compound of formula (/V) that is administered is 3 mg. In some embodiments,
the amount of
the compound of formulat (/V) that is administered is 2.3 mg. In some
embodiments, the
amount of the compound of formula (IHa) that is contained in a capsule is 5.7
mg equivalent
to 4 mg of the compound of formula (/V). In some embodiments, the amount of
the
compound of formula (IHa) that is contained in a capsule is 4.3 mg equivalent
to 3 mg of the
compound of formula (IV). In some embodiments, the amount of the compound of
formula
(IHa) that is contained in a capsule is 3.3 mg equivalent to 2.3 mg of the
compound of
formula (/V).
[0036] In some embodiments, the treatment regimen further comprises
additional
therapeutic agents. In some embodiments, the additional therapeutic agent is
an
immunomodulatory drug. Examples of immunomodulatory drugs include lenalidomide
and
pomalidomide. In some embodiments, the immunomodulatory drug is lenalidomide.
In some
embodiments, the additional therapeutic agent is a steroid. Examples of
steroids include
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dexamethasone and prednisone. In some embodiments, the additional therapeutic
agent is
dexamethasone. In some embodiments, the additional therapeutic agents are
lenalidomide
and dexamethasone.
[0037] In some embodiments, the patient with multiple myeloma has newly
diagnosed
multiple myeloma. In some embodiments, the patient with multiple myeloma has
relapsed
and/or refractory multiple myeloma. In some embodiments, the patient with
multiple
myeloma has relapsed multiple myeloma. In some embodiments, the patient with
multiple
myeloma has refractory myeloma.
[0038] The initiation and progression of multiple myeloma may be influenced
by multiple
mutations in different pathways and genes of the plasma cell. There can be
multiple generic
events that lead to disease progression and treatment resistant disease.
Primary genetic events
include IgH translocations and hyperdiploidy, while secondary genetic events
include copy
number abnormalities, DNA hypomethylation and acquired mutations. Biran et
al., Risk
Stratification in Multiple Myeloma, Part 1, Characterization of High-Risk
Disease,
Hematology and Oncology, 11, Aug 2013 (8) page 489-503.
[0039] Guidelines from the International Myeloma Working Group support a
comprehensive cytogenetic evaluation in all patients at the time of diagnosis
and at relapse.
These recommendations include interphase FISH (Fluorescent In Situ
Hybridization) in
purified plasma cells or in combination with immunofluorescent detection of
light-chain-
restricted plasma cells (cIg-FISH).
[0040] In multiple myeloma, cytogenetic alterations that can be detected by
FISH include,
but are not limited to, t(4:14), t(14:16) and del (17).
Del (17p) =
[0041] The majority of chromosome 17 deletions are hemizygous or impact the
whole p arm.
This genetic event is observed in approximately 10% of new myeloma cases with
the
frequency increasing in later disease stages (Fonseca et al., Blood. 2003 Jun
1;101(11):4569-
75; Tiedemann et al., Leukemia (2008) 22, 1044-1052). The relevant gene
deregulated in
del(17p) is thought to be the tumor suppressor gene TP53, as GEP has shown
that myeloma
samples with monoallelic 17p deletions express significantly less TP53
compared to non
deleted samples (Walker et al., Blood. 2010 Oct 14;116(15):e56-65). Thus in
some cells, the
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CA 02915105 2015-12-10
TP53 gene can be deleted as a result of del 17p deletion. In cases without
del(17p) the rate of
TP53 mutation is <1%, whereas in cases with del(17p) this rises to 25-37%
(Lode et al.,
Haematologica 11/2010; 95(11):1973-6); a finding providing some evidence that
monoallelic
17p deletion contributes to disruption of the remaining allele. The TP53 gene
has been
mapped to 17p13 and is known to function as a transcriptional regulator
influencing cell cycle
arrest, DNA repair, and apoptosis in response to DNA damage. Del(17p) is the
most
important molecular finding for prognostication in multiple myeloma as it is
linked to an
aggressive disease phenotype, a greater degree of extramedullary disease, and
shortened
survival (Fonseca et al., Blood. 2003 Jun 1;101(11):4569-75, Avet-Loiseau et
al., Blood. 2007
Apr 15;109(8):3489-95).
t(4:14) and t(14:16)
[0042] IGH translocations are detectable in approximately 40% of multiple
myeloma
(MM) patients. Most of the translocations detected in MM lead to alteration of
the cell cycle
control, which is a unifying event in early pathogenesis of MM (Bergsagel et
al., Blood,
2005;106(1):296-303). The t(4:14) translocation is detected in about 15% of
patients. T(4:14)
leads to upregulation of two genes MMSET and FGFR3, a receptor tyrosine
kinase. MMSET
protein has histone methylation activity and among the several genes effected
by its activity
there are several cell cycle ¨related genes, such as Cyclin E2, E2F2, Tp53INP1
and CDC25A.
100431 The translocation t(14:16) is seen in about 6% of multiple myeloma
patients and
has also been associated with unfavorable prognosis in multiple myeloma. It
contains the
MAF transcription factor family, which is upregulated as a result of this
translocation.
(Bergsagel et al., Blood. 2005;106(1):296-303).
100441 In some embodiments, the cytogenetic alteration is a deletion in
chromosome 17.
In some embodiments, the cytogenetic alteration is del (17). As used herein,
the terms del
(17) and del (17p) are used interchangeably. In some embodiments, the
cytogenetic alteration
is del (17p13). In some embodiments, there is at least one other cytogenetic
alteration. In
some embodiments, the at least one other cytogenetic alteration is t(4:14). In
some
embodiments, the at least one other cytogenetic alteration is t(14:16). In
some embodiments,
there is at least two other cytogenetic alterations. In some embodiments, the
at least two other
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CA 02915105 2015-12-10
cytogenetic alterations are t(4:14) and t(14:16). In some embodiments, the
cytogenetic
alternation is at least one of del(17), t(4:14), or t((14:16).
[0045] In some embodiments, the cytogenetic alterations are del (17) and
t(4:14). In some
embodiments, the cytogenetic alternations are del (17) and t(14:16). In some
embodiments,
the cytogenetic alterations are del (17), t(4:14) and t(14:16). In some
embodiments, the
cytogenetic alteration is t(4:14).
[0046] In some embodiments, the present disclosure relates to methods of
treating a
patient having multiple myeloma, comprising:
i) selecting a patient based upon the patient having a cytogenetic
alteration at
chromosome 17; and
ii) ii) administering to the patient a treatment regimen comprising a
compound
of formula (Ia) or a pharmaceutically acceptable salt, stereoisomeric or
tautomeric form thereof, wherein Z1 and Z2 are each independently
hydroxyl; or Z1 and Z2 together form a cyclic boronic ester having 2-20
carbon atoms, and optionally one or more heteroatoms selected from N, S,
or O.
[0047] In some embodiments, the present disclosure relates to methods of
evaluating a
patient having multiple myeloma for responsiveness to a treatment regimen
comprising a
compound of formula (Ia) or a pharmaceutically acceptable salt, stereoisomeric
or tautomeric
form thereof, wherein:Z1 and Z2 are each independently hydroxyl; or Z1 and Z2
together form
a cyclic boronic ester having 2-20 carbon atoms, and optionally one or more
heteroatoms
selected from N, S, or 0; comprising the steps:
i) determining whether a patient has a cytogenetic alteration at chromosome
17;
ii) recording the presence or absence of a cytogenetic alteration at
chromosome 17,
and
iii) determining, recommending or selecting an appropriate treatment regimen
based
upon the presence or absence of a cytogenetic alteration at chromosome 17.
[0048] In some embodiments, step iii) comprises determining whether to
begin or
continue the treatment regimen comprising the compound of formula (Ia) or a
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CA 02915105 2015-12-10
pharmaceutically acceptable salt, stereoisomeric or tautomeric form thereof,
based upon the
presence of a cytogenetic alteration at chromosome 17.
[0049] The identification of a correlation between a cytogenetic alteration
that a patient
with multiple myeloma has and clinical responsiveness to a treatment regimen
comprising a
compound of formula (Ia), can be the basis for designing a diagnostic method
to predict those
individuals who will respond to a treatment regimen compromising a compound of
formula
(Ia). Alternatively, such methods can also be used to predict individuals who
will respond to a
treatment regimen comprising a compound of formula (Ia) versus a treatment
regimen not
comprising a compound of formula (Ia).
[0050] The methods for evaluating a patient having multiple myeloma for
responsiveness
or non-responsiveness to a treatment regimen comprising a compound of formula
(Ia) can
include an additional step of recording the value of a parameter related to
the patient's
cytogenetic alteration status.
[0051] "Recording" as used herein, refers to the act or process of making a
record capable
of being accessed or referenced at a later date. In one embodiment, the record
is made in
writing. In one embodiment, the record is made on paper (e.g., written in a
patient's medical
record or written on a batch record), or the record is made in an electronic
medium (e.g., the
record is entered into a computer, for example, the record is entered into an
electronic version
of the patient's medical record or the record is entered into a database). In
another
embodiment, the record is made vocally by recording one's voice. In one
embodiment, the
voice recording is made on, for example, a tape or compact disk. In one
embodiment, the
recorded information contains reference standard value(s).
[0052] The methods for evaluating a patient having multiple myeloma for
responsiveness
or non-responsiveness to a treatment regimen comprising a compound of formula
(Ia) can
include a further step of determining, recommending or selecting an
appropriate treatment
regimen.
[0053] As used herein, "determining an appropriate treatment regimen"
refers to the act or
process of reviewing a patient's genotype; and, optionally, reviewing the
patient's medical
history (e.g., for allergies or intolerances to certain types of drugs, or for
drug
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CA 02915105 2015-12-10
incompatibilities) and assessing the likelihood that the patient will be
responsive to a given
treatment regimen.
[0054] As used herein, "recommending an appropriate treatment regimen"
refers to the
act or process of suggesting, for example, to the patient, to a family member
or caregiver of
the patient, or to medical personnel (e.g., the patient's primary care
physician), a treatment
regimen which is perceived as being favorable for the patient. As used herein,
a
recommendation can be a written or a verbal recommendation.
[0055] As used herein, "selecting an appropriate treatment regimen" refers
to the act or
process of picking or choosing a treatment regimen from other treatment
regimen options for
a patient. In one embodiment, the selection is made upon review of i) the
patient's genotype;
and/or ii) the patient's medical history (e.g., for allergies or intolerances
to certain types of
drugs, for drug incompatibilities and for treatment history) and assessing the
likelihood that
the patient will be responsive to a given treatment regimen. In another
embodiment, the
selection is made based upon a recommendation.
[0056] In some embodimentsõ the present disclosure provides a method of
treating a
patient having multiple myeloma, comprising:
(iii) determining if the patient has a cytogenetic alteration at
cytogenetic alteration
t(4:14); and
(iv) if the patient has a cytogenetic alteration t(4:14), then
administering to a patient
in need thereof a treatment regimen comprising a compound of formula (/):
0 Z1
oNr'N.!Bz2
H "
0
Y (I)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein
ring A is
-14-

CA 02915105 2015-12-10
µ F
0 CI * \ * F 0 'it * Br, \ F
F \ 10
, F CI
CI
Itt.
F . 11,_
Cl 0 µ
*
(00 11/4 0 lk
F F CI, CI CI F F CI
/ / / / / /
CI
1:61
CI CI F F Cl F Cl CICI Or
CI s 114.
CI ;and .
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
100571 In some
embodiments, the present disclosure provides a method of treating a
patient having multiple myeloma, comprising:
(i) determining if the patient has a cytogenetic alteration t(4:14); and
(ii) if the patient has a cytogenetic alteration at t(4:14), then
administering to a
patient in need thereof a treatment regimen comprising a compound of formula
(Ia):
CI H 71
. N N .E3z2
H 0
Y
Cl (Ia);
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof,
wherein Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form
a
cyclic boronic ester having 2-20 carbon atoms, and optionally one or more
heteroatoms selected from N, S, or O.
100581 In some
embodiments, the present disclosure provides a use of a compound of
formula I for treating a patient having multiple myeloma, wherein the use
comprises:
(i) determining if the patient has a cytogenetic alteration at chromosome 17,
and
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CA 02915105 2015-12-10
(ii) if the patient has a cytogenetic alteration at chromosome 17, selecting
the patient
for administration of a treatment regimen comprising a compound of formula
(Ia):
CI
N.rN Bz2
0
CI (Ia)
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
[0059] In some embodiments, the present disclosure relates to methods of
treating a
patient having multiple myeloma, comprising:
i) selecting a patient based upon the patient having a cytogenetic
alteration
t(4:14); and
ii) ii) administering to the patient a treatment regimen comprising a
compound
of formula (la) or a pharmaceutically acceptable salt, stereoisomeric or
tautomeric form thereof, wherein Z1 and Z2 are each independently
hydroxyl; or Z1 and Z2 together form a cyclic boronic ester having 2-20
carbon atoms, and optionally one or more heteroatoms selected from N, S,
or O.
[0060] In some embodiments, the present disclosure relates to use of a
compound of
formula (la) for treating a patient having multiple myeloma, comprising:
i) selecting a patient, based upon the patient having a cytogenetic alteration
at
chromosome 17, for administration of a treatment regimen comprising a compound
of
formula (Ia):
CI
N
0
CI (la);
or a pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein:
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CA 02915105 2015-12-10
Z1 and Z2 are each independently hydroxyl; or Z1 and Z2 together form a cyclic
boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected from
N, S, or O.
[0061] In some embodiments, the present disclosure relates to methods of
evaluating a
patient having multiple myeloma for responsiveness to a treatment regimen
comprising a
compound of formula (Ia) or a pharmaceutically acceptable salt, stereoisomeric
or tautomeric
form thereof, wherein:Z1 and Z2 are each independently hydroxyl; or Z1 and Z2
together form
a cyclic boronic ester having 2-20 carbon atoms, and optionally one or more
heteroatoms
selected from N, S, or 0 comprising the steps:
i) determining whether a patient has a cytogenetic alteration t(4:14);
ii) recording the presence or absence of a cytogenetic alteration t(4:14), and
iii) determining, recommending or selecting an appropriate treatment regimen
based
upon the presence or absence of a cytogenetic alteration t(4:14).
[0062] In some embodiments, step iii) comprises determining whether to
begin or
continue the treatment regimen comprising the compound of formula (Ia) or a
pharmaceutically acceptable salt, stereoisomeric or tautomeric form thereof,
based upon the
presence of a cytogenetic alteration t(4:14).
[0063] In some embodiments, determining if a patient has a cytogenetic
alteration
comprises the steps of:
(i) providing a bone marrow aspirate sample from the patient;
(ii) isolating CD138 positive plasma cells from the sample; and
(iii) performing FISH analysis on the CD138 enriched positive plasma cells.
100641 In some embodiments, the sample is a bone marrow aspirate sample. In
some
embodiments, the sample is blood. In some embodiments, the CD138 positive
plasma cells
are isolated from the sample using fluorescence activated cell sorting. In
some embodiments,
the CD138 positive plasma cells are isolated from the sample using magnetic
activated cell
sorting (MACS). The magnetic or immunomagnetic beads are available from a
number of
commercial sources including Miltenyi Biotec (CA, USA) or Stem Cell
Technologies
(Vancouver, Canada).
[0065] A general principle of prognostic assays involves preparing a sample
or reaction
mixture that may contain a marker, and a probe, under appropriate conditions
and for a time
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CA 02915105 2015-12-10
sufficient to allow the marker and probe to interact and bind, thus forming a
complex that can
be removed and/or detected in the reaction mixture. These assays can be
conducted in a
variety of ways. For example, one method to conduct such an assay would
involve anchoring
the marker or probe onto a solid phase support, also referred to as a
substrate, and detecting
target marker/probe complexes anchored on the solid phase at the end of the
reaction. In one
embodiment of such a method, a sample from a subject, which is to be assayed
for presence
and/or concentration of marker, can be anchored onto a carrier or solid phase
support. In
another embodiment, the reverse situation is possible, in which the probe can
be anchored to a
solid phase and a sample from a subject can be allowed to react as an
unanchored component
of the assay. One example of such an embodiment includes use of an array or
chip which
contains a predictive marker or marker set anchored for expression analysis of
the sample.
100661 There are many established methods for anchoring assay components to
a solid
phase. These include, marker or probe molecules which are immobilized through
conjugation
of biotin and streptavidin. Such biotinylated assay components can be prepared
from biotin-
NHS (N-hydroxy-succinimide) using techniques known in the art (e.g.,
biotinylation kit,
Pierce Chemicals, Rockford, IL), and immobilized in the wells of streptavidin-
coated 96 well
plates (Pierce Chemical). In certain embodiments, the surfaces with
immobilized assay
components can be prepared in advance and stored.
100671 Other suitable carriers or solid phase supports for such assays
include any material
capable of binding the class of molecule to which the marker or probe belongs.
Examples of
supports or carriers include, but are not limited to, glass, polystyrene,
nylon, polypropylene,
nylon, polyethylene, dextran, amylases, natural and modified celluloses,
polyacrylamides,
gabbros, and magnetite. One skilled in the art may know many other suitable
carriers for
binding antibody or antigen, and will be able to adapt such support for use
with the present
disclosure. For example, protein isolated from cells can be run on a
polyacrylamide gel
electrophoresis and immobilized onto a solid phase support such as
nitrocellulose. The
support can then be washed with suitable buffers followed by treatment with
the detectably
labeled antibody. The solid phase support can then be washed with the buffer a
second time
to remove unbound antibody. The amount of bound label on the solid support can
then be
detected by conventional means.
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CA 02915105 2015-12-10
[0068] In order to conduct assays with the above mentioned approaches, the
non-
immobilized component is added to the solid phase upon which the second
component is
anchored. After the reaction is complete, uncomplexed components may be
removed (e.g., by
washing) under conditions such that any complexes formed will remain
immobilized upon the
solid phase. The detection of marker/probe complexes anchored to the solid
phase can be
accomplished in a number of methods outlined herein.
[0069] In an embodiment, the probe, when it is the unanchored assay
component, can be
labeled for the purpose of detection and readout of the assay, either directly
or indirectly, with
detectable labels discussed herein and which are well-known to one skilled in
the art. The
term "labeled", with regard to the probe (e.g., nucleic acid or antibody), is
intended to
encompass direct labeling of the probe by coupling (i.e., physically linking)
a detectable
substance to the probe, as well as indirect labeling of the probe by
reactivity with another
reagent that is directly labeled. An example of indirect labeling includes
detection of a
primary antibody using a fluorescently labeled secondary antibody. It is also
possible to
directly detect marker/probe complex formation without further manipulation or
labeling of
either component (marker or probe), for example by utilizing the technique of
fluorescence
energy transfer (FET, see, for example, Lakowicz et al., U.S. Patent No.
5,631,169;
Stavrianopoulos, et al., U.S. Patent No. 4,868,103). A fluorophore label on
the first, 'donor'
molecule is selected such that, upon excitation with incident light of
appropriate wavelength,
its emitted fluorescent energy will be absorbed by a fluorescent label on a
second 'acceptor'
molecule, which in turn is able to fluoresce due to the absorbed energy.
Alternately, the
'donor' protein molecule may simply utilize the natural fluorescent energy of
tryptophan
residues. Labels are chosen that emit different wavelengths of light, such
that the 'acceptor'
molecule label may be differentiated from that of the 'donor'. Since the
efficiency of energy
transfer between the labels is related to the distance separating the
molecules, spatial
relationships between the molecules can be assessed. In a situation in which
binding occurs
between the molecules, the fluorescent emission of the 'acceptor' molecule
label in the assay
should be maximal. An FET binding event can be conveniently measured through
standard
fluorometric detection means well known in the art (e.g., using a
fluorimeter).
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CA 02915105 2015-12-10
[0070] As used herein, the term "hybridizes" is intended to describe
conditions for
hybridization and washing under which nucleotide sequences that are
significantly identical or
homologous to each other remain hybridized to each other. In some embodiments,
the
conditions are such that sequences at least about 70%, at least about 80%, at
least about 85%,
90% or 95% identical to each other remain hybridized to each other for
subsequent
amplification and/or detection. Stringent conditions vary according to the
length of the
involved nucleotide sequence but are known to those skilled in the art and can
be found or
determined based on teachings in Current Protocols in Molecular Biology,
Ausubel et al.,
eds., John Wiley & Sons, Inc. (1995), sections 2, 4 and 6. Additional
stringent conditions
and formulas for determining such conditions can be found in Molecular
Cloning: A
Laboratory Manual, Sambrook et al., Cold Spring Harbor Press, Cold Spring
Harbor, NY
(1989), chapters 7, 9 and 11. A non-limiting example of stringent
hybridization conditions for
hybrids that are at least 10 basepairs in length includes hybridization in 4X
sodium
chloride/sodium citrate (SSC), at about 65-70 C (or hybridization in 4X SSC
plus 50%
formamide at about 42-50 C) followed by one or more washes in 1X SSC, at about
65-70 C.
A non-limiting example of highly stringent hybridization conditions for such
hybrids includes
hybridization in 1X SSC, at about 65-70 C (or hybridization in 1X SSC plus 50%
formamide
at about 42-50 C) followed by one or more washes in 0.3X SSC, at about 65-70
C. A non-
limiting example of reduced stringency hybridization conditions for such
hybrids includes
hybridization in 4X SSC, at about 50-60 C (or alternatively hybridization in
6X SSC plus
50% formamide at about 40-45 C) followed by one or more washes in 2X SSC, at
about 50-
60 C. Ranges intermediate to the above-recited values, e.g., at 65-70 C or at
42-50 C are
also intended to be encompassed by the present disclosure. Another example of
stringent
hybridization conditions are hybridization in 6X sodium chloride/sodium
citrate (SSC) at
about 45 C, followed by one or more washes in 0.2X SSC, 0.1% SDS at 50-65 C. A
further
example of stringent hybridization buffer is hybridization in 1 M NaC1, 50 mM
2-(N-
morpholino) ethanesulfonic acid (MES) buffer (pH 6.5), 0.5% sodium sarcosine
and 30%
formamide. SSPE (1xSSPE is 0.15M NaC1, 10mM NaH2PO4, and 1.25mM EDTA, pH 7.4)
can be substituted for SSC (1xSSC is 0.15M NaC1 and 15mM sodium citrate) in
the
hybridization and wash buffers; washes are performed for 15 minutes each after
hybridization
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CA 02915105 2015-12-10
is complete The hybridization temperature for hybrids anticipated to be less
than 50 base pairs
in length should be 5-10 C less than the melting temperature (Tm) of the
hybrid, where Tm is
determined according to the following equations. For hybrids less than 18 base
pairs in
length, Tm( C) = 2(# of A + T bases) + 4(# of G + C bases). For hybrids
between 18 and 49
base pairs in length, Tm( C) = 81.5 + 16.6(logio[Na]) + 0.41(%G+C) - (600/N),
where N is
the number of bases in the hybrid, and [Na] is the concentration of sodium
ions in the
hybridization buffer ([Na] for 1xSSC = 0.165 M). It will also be recognized by
the skilled
practitioner that additional reagents may be added to hybridization and/or
wash buffers to
decrease non-specific hybridization of nucleic acid molecules to membranes,
for example,
nitrocellulose or nylon membranes, including but not limited to blocking
agents (e.g., BSA or
salmon or herring sperm carrier DNA), detergents (e.g., SDS), chelating agents
(e.g., EDTA),
Ficoll, polyvinylpyrrolidone (PVP) and the like. When using nylon membranes,
in particular,
an additional non-limiting example of stringent hybridization conditions is
hybridization in
0.25-0.5M NaH2PO4, 7% SDS at about 65 C, followed by one or more washes at
0.02M
NaH2PO4, 1% SDS at 65 C, see e.g., Church and Gilbert (1984) Proc. Natl. Acad.
Sci. USA
81:1991-1995, (or alternatively 0.2X SSC, 1% SDS). A primer or nucleic acid
probe can be
used alone in a detection method, or a primer can be used together with at
least one other
primer or nucleic acid probe in a detection method. Primers can also be used
to amplify at
least a portion of a nucleic acid. Nucleic acid probes of the disclosure refer
to nucleic acids
which hybridize to the region of interest and which are not further extended.
For example, a
nucleic acid probe is a nucleic acid which specifically hybridizes to a mutant
region of a
biomarker, and which by hybridization or absence of hybridization to the DNA
of a patient or
the type of hybrid formed can be indicative of the presence or identity of the
mutation of the
biomarker or the amount of marker activity.
100711 In one format, the RNA is immobilized on a solid surface and
contacted with a
probe, for example by running the isolated RNA on an agarose gel and
transferring the RNA
from the gel to a membrane, such as nitrocellulose. In an alternative format,
the nucleic acid
probe(s) are immobilized on a solid surface and the RNA is contacted with the
probe(s), for
example, in an AFFYMETRIX gene chip array or a SNP chip (Santa Clara, CA) or
customized array using a marker set comprising at least one marker indicative
of treatment
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CA 02915105 2015-12-10
outcome. A skilled artisan can readily adapt known RNA and DNA detection
methods for
use in detecting the amount of the markers of the present disclosure. For
example, the high
density microarray or branched DNA assay can benefit from a higher
concentration of tumor
cell in the sample, such as a sample which had been modified to isolate tumor
cells as
described in earlier sections. In a related embodiment, a mixture of
transcribed
polynucleotides obtained from the sample is contacted with a substrate having
fixed thereto a
polynucleotide complementary to or homologous with at least a portion (e.g.,
at least 7, 10,
15, 20, 25, 30, 40, 50, 100, 500, or more nucleotide residues or 10 to 50, 15
to 40 or 15 to 30
consecutive nucleotides) of a marker nucleic acid. If polynucleotides
complementary to or
homologous with the marker are differentially detectable on the substrate
(e.g., detectable
using different chromophores or fluorophores, or fixed to different selected
positions), then
the levels of expression of a plurality of markers can be assessed
simultaneously using a
single substrate (e.g., a "gene chip" microarray of polynucleotides fixed at
selected positions).
In an embodiment when a method of assessing marker expression is used which
involves
hybridization of one nucleic acid with another, the hybridization can be
performed under
stringent hybridization conditions.
[0072] In an embodiment, a mutation in a marker can be identified by
sequencing a
nucleic acid, e.g., a DNA, RNA, cDNA, genomic DNA or a protein correlated with
the
marker gene. There are several sequencing methods known in the art to sequence
nucleic
acids. A primer can be designed to bind to a region comprising a potential
mutation site or
can be designed to complement the mutated sequence rather than the wild type
sequence.
Primer pairs can be designed to bracket a region comprising a potential
mutation in a marker
gene. A primer or primer pair can be used for sequencing one or both strands
of DNA
corresponding to the marker gene. A primer can be used in conjunction with a
probe to
amplify a region of interest prior to sequencing to boost sequence amounts for
detection of a
mutation in a marker gene. Examples of regions which can be sequenced include
an entire
gene, transcripts of the gene and a fragment of the gene or the transcript,
e.g., one or more of
exons or untranslated regions. Examples of mutations to target for primer
selection and
sequence or composition analysis can be found in public databases which
collect mutation
information, such as COSMIC and dbGaP.
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[0073] In an embodiment, DNA, e.g., genomic DNA corresponding to the wild
type or
mutated marker can be analyzed both by in situ and by in vitro formats in a
biological sample
using methods known in the art. DNA can be directly isolated from the sample
or isolated
after isolating another cellular component, e.g., RNA or protein. Kits are
available for DNA
isolation, e.g., QIAAMP DNA Micro Kit (Qiagen, Valencia, CA). DNA also can be

amplified using such kits.
[0074] In one embodiment, expression of a marker is assessed by preparing
mRNA/cDNA (i.e., a transcribed polynucleotide) from cells in a patient sample,
and by
hybridizing the mRNA/cDNA with a reference polynucleotide which is a
complement of a
marker nucleic acid, or a fragment thereof. cDNA can, optionally, be amplified
using any of
a variety of polymerase chain reaction methods prior to hybridization with the
reference
polynucleotide. Expression of one or more markers likewise can be detected
using
quantitative PCR to assess the level of expression of the marker(s). An
example of the use of
measuring mRNA levels is that an inactivating mutation in a marker gene can
result in an
altered level of mRNA in a cell. The level can be upregulated due to feedback
signaling
protein production in view of nonfunctional or absent protein or downregulated
due to
instability of an altered mRNA sequence. Alternatively, any of the many known
methods of
detecting mutations or variants (e.g. single nucleotide polymorphisms,
deletions, etc.,
discussed above) of a marker of the disclosure may be used to detect
occurrence of a mutation
in a marker gene in a patient.
[0075] An example of direct measurement is quantification of transcripts.
As used herein,
the level or amount of expression refers to the absolute amount of expression
of an mRNA
encoded by the marker or the absolute amount of expression of the protein
encoded by the
marker. As an alternative to making determinations based on the absolute
expression amount
of selected markers, determinations may be based on normalized expression
amounts.
Expression amount can be normalized by correcting the absolute expression
level of a marker
upon comparing its expression to the expression of a control marker that is
not a marker, e.g.,
in a housekeeping role that is constitutively expressed. Suitable markers for
normalization
also include housekeeping genes, such as the actin gene or beta-2
microglobulin. Reference
markers for data normalization purposes include markers which are ubiquitously
expressed
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CA 02915105 2015-12-10
and/or whose expression is not regulated by oncogenes. Constitutively
expressed genes are
known in the art and can be identified and selected according to the relevant
tissue and/or
situation of the patient and the analysis methods. Such normalization allows
one to compare
the expression level in one sample, to another sample, e.g., between samples
from different
times or different subjects. Further, the expression level can be provided as
a relative
expression level. The baseline of a genomic DNA sample, e.g., diploid copy
number, can be
determined by measuring amounts in cells from subjects without a tumor or in
non-tumor
cells from the patient. To determine a relative amount of a marker or marker
set, the amount
of the marker or marker set is determined for at least 1, or 2, 3, 4, 5, or
more samples, e.g., 7,
10, 15, 20 or 50 or more samples in order to establish a baseline, prior to
the determination of
the expression level for the sample in question. To establish a baseline
measurement, the
mean amount or level of each of the markers or marker sets assayed in the
larger number of
samples is determined and this is used as a baseline expression level for the
biomarkers or
biomarker sets in question. The amount of the marker or marker set determined
for the test
sample (e.g., absolute level of expression) is then divided by the baseline
value obtained for
that marker or marker set. This provides a relative amount and aids in
identifying abnormal
levels of marker protein activity.
100761 Probes based on the sequence of a nucleic acid molecule of the
disclosure can be
used to detect transcripts or genomic sequences corresponding to one or more
markers of the
disclosure. The probe can comprise a label group attached thereto, e.g., a
radioisotope, a
fluorescent compound, an enzyme, or an enzyme co-factor. Such probes can be
used as part
of a diagnostic test kit for identifying cells or tissues which express the
protein, such as by
measuring levels of a nucleic acid molecule encoding the protein in a sample
of cells from a
subject, e.g., detecting mRNA levels or determining whether a gene encoding
the protein has
been mutated or deleted.
[0077] Primers or nucleic acid probes comprise a nucleotide sequence
complementary to a
specific a marker or a mutated region thereof and are of sufficient length to
selectively
hybridize with a marker gene or nucleic acid associated with a marker gene.
Primers and
probes can be used to aid in the isolation and sequencing of marker nucleic
acids. In one
embodiment, the primer or nucleic acid probe, e.g., a substantially purified
oligonucleotide,
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CA 02915105 2015-12-10
comprises a region having a nucleotide sequence which hybridizes under
stringent conditions
to about 6, 8, 10, 12, or 15, 20, 25, 30, 40, 50, 60, 75, 100 or more
consecutive nucleotides of
a marker gene. For example, a primer or nucleic acid probe comprising a
nucleotide sequence
of at least about 15 consecutive nucleotides, at least about 25 nucleotides or
having from
about 15 to about 20 nucleotides, 10 to 50 consecutive nucleotides, 12 to 35
consecutive
nucleotides, 15 to 50 consecutive nucleotides, 20 to 100 consecutive
nucleotides. Nucleic
acid analogs can be used as binding sites for hybridization. An example of a
suitable nucleic
acid analogue is peptide nucleic acid (see, e.g., Egholm et al., Nature
363:566 568 (1993);
U.S. Pat. No. 5,539,083).
100781 Primers or nucleic acid probes can be selected using an algorithm
that takes into
account binding energies, base composition, sequence complexity, cross-
hybridization
binding energies, and secondary structure (see Friend et al., International
Patent Publication
WO 01/05935, published Jan. 25, 2001; Hughes et al., Nat. Biotech. 19:342-7
(2001). Useful
primers or nucleic acid probes of the disclosure bind sequences which are
unique for each
transcript, e.g., target mutated regions and can be used in PCR for
amplifying, detecting and
sequencing only that particular nucleic acid, e.g., transcript or mutated
transcript. One of skill
in the art can design primers and nucleic acid probes for the markers
disclosed herein or
related markers with similar characteristics, e.g., markers on the chromosome
loci, or
mutations in different regions of the same marker gene described herein, using
the skill in the
art, e.g., adjusting the potential for primer or nucleic acid probe binding to
standard
sequences, mutants or allelic variants by manipulating degeneracy or GC
content in the
primer or nucleic acid probe. Computer programs that are well known in the art
are useful in
the design of primers with the required specificity and optimal amplification
properties, such
as Oligo version 5.0 (National Biosciences, Plymouth, MN). While perfectly
complementary
nucleic acid probes and primers can be used for detecting the markers
described herein and
mutants, polymorphisms or alleles thereof, departures from complete
complementarity are
contemplated where such departures do not prevent the molecule from
specifically
hybridizing to the target region. For example, an oligonucleotide primer may
have a non-
complementary fragment at its 5' end, with the remainder of the primer being
complementary
to the target region. Alternatively, non-complementary nucleotides may be
interspersed into
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CA 02915105 2015-12-10
the nucleic acid probe or primer as long as the resulting probe or primer is
still capable of
specifically hybridizing to the target region.
[0079] FISH (fluorescent in situ hybridization) is a cytogenetic technique
developed by
biomedical researchers in the early 1980s that is used to detect and localize
the presence or
absence of specific DNA sequences on chromosomes. See Speicher and Carter,
Nat. Rev.
Genet.2005 Oct; 6(10) 782-792. FISH uses fluorescent probes that bind to only
those parts of
the chromosome with which they show a high degree of sequence complementarily.
The
IMWG recommends FISH testing for multiple myeloma patients on nuclei from
purified
plasma cells; Shi et al., Acta Medica International, Jul-Dec 2015 2(2), page
168-174.
[0080] Probes for the cytogenetic alternations del (17), t(4:14) and
t(14:16) are available
from a variety of commercial sources, including, for example, Empire Genomics,
(Buffalo,
NY), Kreatech Inc., (part of Leica Biosystems, Illinois, USA) and Biocare
Medical (CA,
USA). Fluorescence microscopy can be used to find out where the fluorescent
probe is bound
to the chromosomes. Cells are fixed, then permeabilized to allow target
accessibility. A
target-specific probe, e.g., a probe composed of 20 oligonucleotide pairs,
hybridizes to the
target RNA(s). Separate but compatible signal amplification systems enable the
multiplex
assay (more than two targets per assay, such as four, six, eight or more
targets). Signal
amplification is achieved via a series of sequential hybridization steps. At
the end of the assay
the samples are visualized under a fluorescence microscope.
[0081] Preparation and hybridization process: First, a probe is
constructed. The probe
must be large enough to hybridize specifically with its target but not so
large as to impede the
hybridization process. The probe is tagged directly with fluorophores, with
targets for
antibodies or with biotin. Tagging can be done in various ways, such as nick
translation, or
PCR using tagged nucleotides.
[0082] Secondly, an interphase or metaphase chromosome preparation is
produced. The
chromosomes are firmly attached to a substrate, usually glass. Repetitive DNA
sequences
must be blocked by adding short fragments of DNA to the sample. The probe is
then applied
to the chromosome DNA and incubated for approximately 12 hours while
hybridizing.
Several wash steps remove all unhybridized or partially hybridized probes. The
results are
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CA 02915105 2015-12-10
then visualized and quantified using a microscope that is capable of exciting
the dye and
recording images.
[0083] If the fluorescent signal is weak, amplification of the signal may
be necessary in
order to exceed the detection threshold of the microscope. Fluorescent signal
strength depends
on many factors such as probe labeling efficiency, the type of probe, and the
type of dye.
Fluorescently tagged antibodies or streptavidin are bound to the dye molecule.
These
secondary components are selected so that they have a strong signal. One way
to amplify the
fluorescent signal is using tyramide-based signal amplification technology
which employs
horseradish peroxidase (HRP) to generate high-density fluorescence labeling at
the site of
probe hybridization.
100841 Based upon determining a noise level for the commercially available
probe for del
(17) a cut-off value of 5 cells per hundred (5%) was used to classify an
individual as have del
(17). In some embodiments, a cut-off value of 5% classifies an individual as
having del (17).
In some embodiments a cut-off value of 20% classifies an individual as having
del (17). In
some embodiments, a cut-off value of 60% classifies an individual as having
del (17).
Pharmaceutical compositions
100851 The compounds and pharmaceutical compositions disclosed herein may
be
administered by any route, including intradermally, subcutaneously, orally,
intraarterially or
intravenously. In one embodiment, administration will be by the intravenous
route. Parenteral
administration can be provided in a bolus or by infusion.
100861 The concentration of a disclosed compound in a pharmaceutically
acceptable
mixture will vary depending on several factors, including the dosage of the
compound to be
administered, the pharmacokinetic characteristics of the compound(s) employed,
and the route
of administration. The agent may be administered in a single dose or in repeat
doses.
Treatments may be administered daily or more frequently depending upon a
number of
factors, including the overall health of a patient, and the formulation and
route of
administration of the selected compound(s).
[0087] If a pharmaceutically acceptable salt of a compound of the present
disclosure is
utilized in these compositions, the salt preferably is derived from an
inorganic or organic acid
or base. For reviews of suitable salts, see, e.g., Berge et al, J. Pharm. Sci.
66:1-19 (1977) and
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Remington: The Science and Practice of Pharmacy, 20th Ed., ed. A. Gennaro,
Lippincott
Williams & Wilkins, 2000.
[0088] Nonlimiting examples of suitable acid addition salts include the
following: acetate,
adipate, alginate, aspartate, benzoate, benzene sulfonate, bisulfate,
butyrate, citrate,
camphorate, camphor sulfonate, cyclopentanepropionate, digluconate,
dodecylsulfate,
ethanesulfonate, fumarate, lucoheptanoate, glycerophosphate, hemisulfate,
heptanoate,
hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate,
lactate,
maleate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, oxalate,
pamoate, pectinate,
persulfate, 3-phenyl-propionate, picrate, pivalate, propionate, succinate,
tartrate, thiocyanate,
tosylate and undecanoate.
[0089] Suitable base addition salts include, ammonium salts, alkali metal
salts, such as
sodium and potassium salts, alkaline earth metal salts, such as calcium and
magnesium salts,
salts with organic bases, such as dicyclohexylamine, N-methyl-D-glucamine, t-
butylamine,
ethylene diamine, ethanolamine, and choline, and salts with amino acids such
as arginine,
lysine, and so forth.
[0090] Also, basic nitrogen-containing groups may be quaternized with such
agents as
lower alkyl halides, such as methyl, ethyl, propyl, and butyl chlorides,
bromides and iodides;
dialkyl sulfates, such as dimethyl, diethyl, dibutyl and diamyl sulfates, long
chain halides such
as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides,
aralkyl halides, such as
benzyl and phenethyl bromides and others. Water or oil-soluble or dispersible
products are
thereby obtained.
[0091] The term "pharmaceutically acceptable carrier" is used herein to
refer to a material
that is compatible with a recipient subject, preferably a mammal, more
preferably a human,
and is suitable for delivering an active agent to the target site without
terminating the activity
of the agent. The toxicity or adverse effects, if any, associated with the
carrier preferably are
commensurate with a reasonable risk/benefit ratio for the intended use of the
active agent.
[0092] The terms "carrier", "adjuvant", or "vehicle" are used
interchangeably herein, and
include any and all solvents, diluents, and other liquid vehicles, dispersion
or suspension aids,
surface active agents, isotonic agents, thickening or emulsifying agents,
preservatives, solid
binders, lubricants and the like, as suited to the particular dosage form
desired. Remington:
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CA 02915105 2015-12-10
The Science and Practice of Pharmacy, 20th Ed., ed. A. Gennaro, Lippincott
Williams &
Wilkins, 2000 discloses various carriers used in formulating pharmaceutically
acceptable
compositions and known techniques for the preparation thereof. Except insofar
as any
conventional carrier medium is incompatible with the compounds of the
disclosure, such as
by producing any undesirable biological effect or otherwise interacting in a
deleterious
manner with any other component(s) of the pharmaceutically acceptable
composition, its use
is contemplated to be within the scope of this disclosure. Some examples of
materials which
can serve as pharmaceutically acceptable carriers include, but are not limited
to, ion
exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as
human serum
albumin, buffer substances such as disodium hydrogen phosphate, potassium
hydrogen
phosphate, sodium carbonate, sodium bicarbonate, potassium carbonate,
potassium
bicarbonate, magnesium hydroxide and aluminum hydroxide, glycine, sorbic acid,
or
potassium sorbate, partial glyceride mixtures of saturated vegetable fatty
acids, water,
pyrogen-free water, salts or electrolytes such as protamine sulfate, disodium
hydrogen
phosphate, potassium hydrogen phosphate, sodium chloride, and zinc salts,
colloidal silica,
magnesium trisilicate, polyvinyl pyrrolidone, polyacrylates, waxes,
polyethylene-
polyoxypropylene-block polymers, wool fat, sugars such as lactose, glucose,
sucrose, starches
such as corn starch and potato starch, cellulose and its derivatives such as
sodium
carboxymethyl cellulose, ethyl cellulose and cellulose acetate, powdered
tragacanth; malt,
gelatin, talc, excipients such as cocoa butter and suppository waxes, oils
such as peanut oil,
cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean
oil, glycols such as
propylene glycol and polyethylene glycol, esters such as ethyl oleate and
ethyl laurate, agar,
alginic acid, isotonic saline, Ringer's solution, alcohols such as ethanol,
isopropyl alcohol,
hexadecyl alcohol, and glycerol, cyclodextrins, lubricants such as sodium
lauryl sulfate and
magnesium stearate, petroleum hydrocarbons such as mineral oil and petrolatum.
Coloring
agents, releasing agents, coating agents, sweetening, flavoring and perfuming
agents,
preservatives and antioxidants can also be present in the composition,
according to the
judgment of the formulator.
[0093] The pharmaceutical compositions of the disclosure can be
manufactured by
methods known in the art such as conventional granulating, mixing, dissolving,
encapsulating,
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CA 02915105 2015-12-10
lyophilizing, or emulsifying processes, among others. Compositions may be
produced in
various forms, including granules, precipitates, or particulates, powders,
including freeze
dried, rotary dried or spray dried powders, amorphous powders, tablets,
capsules, syrup,
suppositories, injections, emulsions, elixirs, suspensions or solutions.
Formulations may
optionally contain solvents, diluents, and other liquid vehicles, dispersion
or suspension aids,
surface active agents, pH modifiers, isotonic agents, thickening or
emulsifying agents,
stabilizers and preservatives, solid binders, lubricants and the like, as
suited to the particular
dosage form desired.
[0094] According to another embodiment, the compositions of this disclosure
are
formulated for pharmaceutical administration to a mammal, preferably a human
being. Such
pharmaceutical compositions of the present disclosure may be administered
orally,
parenterally, by inhalation spray, topically, rectally, nasally, buccally,
vaginally or via an
implanted reservoir. The term "parenteral" as used herein includes
subcutaneous, intravenous,
intramuscular, intra-articular, intra-synovial, intrasternal, intrathecal,
intrahepatic,
intralesional and intracranial injection or infusion techniques. Preferably,
the compositions are
administered orally, intravenously, or subcutaneously. The formulations of the
disclosure
may be designed to be short-acting, fast-releasing, or long-acting. Still
further, compounds
can be administered in a local rather than systemic means, such as
administration (e.g., by
injection) at a tumor site.
[0095] Liquid dosage forms for oral administration include, but are not
limited to,
pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions,
syrups and
elixirs. In addition to the active compounds, the liquid dosage forms may
contain inert
diluents commonly used in the art such as, for example, water or other
solvents, solubilizing
agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl
carbonate, ethyl acetate,
benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol,
cyclodextrins,
dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ,
olive, castor, and
sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and
fatty acid esters of
sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions
can also include
adjuvants such as wetting agents, emulsifying and suspending agents,
sweetening, flavoring,
and perfuming agents.
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CA 02915105 2015-12-10
[0096] Injectable preparations, for example, sterile injectable aqueous or
oleaginous
suspensions may be formulated according to the known art using suitable
dispersing or
wetting agents and suspending agents. The sterile injectable preparation may
also be a sterile
injectable solution, suspension or emulsion in a nontoxic parenterally
acceptable diluent or
solvent, for example, as a solution in 1,3-butanediol. Among the acceptable
vehicles and
solvents that may be employed are water, Ringer's solution, U.S.P. and
isotonic sodium
chloride solution. In addition, sterile, fixed oils are conventionally
employed as a solvent or
suspending medium. For this purpose any bland fixed oil can be employed
including synthetic
mono- or diglycerides. In addition, fatty acids such as oleic acid are used in
the preparation of
injectables. The injectable formulations can be sterilized, for example, by
filtration through a
bacterial-retaining filter, or by incorporating sterilizing agents in the form
of sterile solid
compositions which can be dissolved or dispersed in sterile water or other
sterile injectable
medium prior to use. Compositions formulated for parenteral administration may
be injected
by bolus injection or by timed push, or may be administered by continuous
infusion.
[0097] Solid dosage forms for oral administration include capsules,
tablets, pills, powders,
and granules. In such solid dosage forms, the active compound is mixed with at
least one
inert, pharmaceutically acceptable excipient or carrier such as sodium citrate
or dicalcium
phosphate and/or a) fillers or extenders such as starches, lactose, sucrose,
glucose, mannitol,
and silicic acid, b) binders such as, for example, carboxyrnethylcellulose,
alginates, gelatin,
polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol,
d) disintegrating
agents such as agar, calcium carbonate, potato or tapioca starch, alginic
acid, certain silicates,
and sodium carbonate, e) solution retarding agents such as paraffin, f)
absorption accelerators
such as quaternary ammonium compounds, g) wetting agents such as, for example,
cetyl
alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite
clay, and i)
lubricants such as talc, calcium stearate, magnesium stearate, solid
polyethylene glycols,
sodium lauryl sulfate, and mixtures thereof In the case of capsules, tablets
and pills, the
dosage form may also comprise buffering agents such as phosphates or
carbonates.
[0098] Solid compositions of a similar type may also be employed as fillers
in soft and
hard-filled gelatin capsules using such excipients as lactose or milk sugar as
well as high
molecular weight polyethylene glycols and the like. The solid dosage forms of
tablets,
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dragees, capsules, pills, and granules can be prepared with coatings and
shells such as enteric
coatings and other coatings well known in the pharmaceutical formulating art.
They may
optionally contain opacifying agents and can also be of a composition that
they release the
active ingredient(s) only, or preferentially, in a certain part of the
intestinal tract, optionally,
in a delayed manner. Examples of embedding compositions that can be used
include
polymeric substances and waxes. Solid compositions of a similar type may also
be employed
as fillers in soft and hard-filled gelatin capsules using such excipients as
lactose or milk sugar
as well as high molecular weight polyethylene glycols and the like.
100991 The active compounds can also be in micro-encapsulated form with one
or more
excipients as noted above. The solid dosage forms of tablets, dragees,
capsules, pills, and
granules can be prepared with coatings and shells such as enteric coatings,
release controlling
coatings and other coatings well known in the pharmaceutical formulating art.
In such solid
dosage forms the active compound may be admixed with at least one inert
diluent such as
sucrose, lactose or starch. Such dosage forms may also comprise, as is normal
practice,
additional substances other than inert diluents, e.g., tableting lubricants
and other tableting
aids such a magnesium stearate and microcrystalline cellulose. In the case of
capsules, tablets
and pills, the dosage forms may also comprise buffering agents. They may
optionally contain
opacifying agents and can also be of a composition that they release the
active ingredient(s)
only, or preferentially, in a certain part of the intestinal tract,
optionally, in a delayed manner.
Examples of embedding compositions that can be used include polymeric
substances and
waxes.
1001001 Dosage forms for topical or transdermal administration of a compound
of this
disclosure include ointments, pastes, creams, lotions, gels, powders,
solutions, sprays,
inhalants or patches. The active component is admixed under sterile conditions
with a
pharmaceutically acceptable carrier and any needed preservatives or buffers as
may be
required. Ophthalmic formulation, ear drops, and eye drops are also
contemplated as being
within the scope of this disclosure. Additionally, the present disclosure
contemplates the use
of transdermal patches, which have the added advantage of providing controlled
delivery of a
compound to the body. Such dosage forms can be made by dissolving or
dispensing the
compound in the proper medium. Absorption enhancers can also be used to
increase the flux
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CA 02915105 2015-12-10
of the compound across the skin. The rate can be controlled by either
providing a rate
controlling membrane or by dispersing the compound in a polymer matrix or gel.
[00101] Compositions for use in the method of the disclosure may be formulated
in unit
dosage form for ease of administration and uniformity of dosage. The
expression "unit
dosage form" as used herein refers to a physically discrete unit of agent
appropriate for the
patient to be treated. It will be understood, however, that the total daily
usage of the
compounds and compositions of the present disclosure will be decided by the
attending
physician within the scope of sound medical judgment. A unit dosage form for
parenteral
administration may be in ampoules or in multi-dose containers.
Articles of manufacture
[00102] In some embodiments, the present disclosure relates to an article of
manufacture
comprising:
i) a compound of formula (Ia) or a pharmaceutically acceptable salt,
stereoisomeric or tautomeric form thereof, wherein Z1 and Z2 are each
independently hydroxyl; or Z1 and Z2 together form a cyclic boronic ester
having 2-20 carbon atoms, and optionally one or more heteroatoms selected
from N, S, or 0; and
ii) instructions for determining the appropriateness of use of said
composition by
determining whether a patient has a cytogenetic alteration at chromosome 17.
[00103] In some embodiments, the present disclosure relates to an article of
manufacture
comprising:
i) reagents for making a determination whether a patient has a cytogenetic
alteration at chromosome 17, and
ii) instructions for determining the appropriateness of use of a
pharmaceutical
composition comprising a compound of formula (Ia) or a pharmaceutically
acceptable salt, stereoisomeric or tautomeric form thereof, wherein Z1 and Z2
are
each independently hydroxyl; or Z1 and Z2 together form a cyclic boronic ester

having 2-20 carbon atoms, and optionally one or more heteroatoms selected from

N, S, or 0 on the basis of said determination.
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CA 02915105 2015-12-10
[00104] In some embodiments, the present disclosure relates to an article of
manufacture
comprising:
i) a pharmaceutical composition comprising a compound of formula (Ia) or a
pharmaceutically acceptable salt, stereoisomeric or tautomeric form
thereof, wherein Z1 and Z2 are each independently hydroxyl; or Z1 and Z2
together form a cyclic boronic ester having 2-20 carbon atoms, and
optionally one or more heteroatoms selected from N, S, or 0;
ii) reagents for making a determination whether a patient has a cytogenetic

alteration at chromosome 17; and
iii) instructions for determining the appropriateness of use of a
pharmaceutical
composition comprising a compound of formula (Ia) or a pharmaceutically
acceptable salt, stereoisomeric or tautomeric form on the basis of said
deteimination.
[00105] In some embodiments, the present disclosure relates to the use of a
reagent to
determine whether a patient has a cytogenetic alteration at chromosome 17, in
the
manufacture of a kit for determining the appropriateness of use of a
pharmaceutical
composition comprising a compound of formula (Ia) or a pharmaceutically
acceptable salt,
stereoisomeric or tautomeric form thereof, wherein Z1 and Z2 are each
independently
hydroxyl; or Z1 and Z2 together form a cyclic boronic ester having 2-20 carbon
atoms, and
optionally one or more heteroatoms selected from N, S, or 0 on the basis of
said
determination; comprising identifying whether the patient has the cytogenetic
alteration and
determining to treat the patient with the compound if the patient has the
cytogenetic alteration.
EXAMPLES
C16010 Clinical Trial
[00106] The study is an international, randomized, double-blind, placebo
controlled
clinical trial designed to compare the efficacy and safety of two treatment
regimens
administered until disease progression or unacceptable toxicity; ixazomib plus
lenalidomide
and dexamethasone versus placebo plus lenalidomide and dexamethasone in adult
patients
with relapsed and/or refractory multiple myeloma.
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CA 02915105 2015-12-10
[00107] Subjects included in the study have a confirmed diagnosis of multiple
myeloma,
have received one to three prior therapies and met other eligibility criteria.
Patients who are
refractory to prior lenalidomide or proteasome inhibitor-based therapy are
excluded. The
primary endpoint is progression free survival (PFS) as assessed by an
independent review
committee blinded to treatment, per IMWG criteria.
[00108] Table 1 below provides information on the conditions and interventions
for each
patient group.
Table 1:
Arms Assigned Interventions
Active Drug: ixazomib + lenalidomide + dexamethasone
Comparator: Patients will receive single oral dose of ixazomib (4.0mg) on
days 1,8,15
ixazomib + and single oral dose of Lenalidomide (25mg) on days 1-21 and
single oral
lenalidomide + dose of Dexamethasone (40mg) on days 1,8, 15 and 22 every 28
days
dexamethasone until disease progression
Placebo Drug: Placebo + Lenalidomide + Dexamethasone
Comparator: Patients will receive single oral dose of Placebo on days
1,8,15 and single
placebo + oral dose of Lenalidomide (25mg) on days 1-21 and single oral
dose of
lenalidomide + Dexamethasone (40mg) on days 1,8, 15 and 22 every 28 days until

dexamethasone disease progression
Sample processing:
1001091 Bone marrow samples are received, CD138 positive plasma cells are
isolated using magnetic bead sorting techniques. 10,000 CD138 positive
enriched
plasma cells are then used for cytogenetic testing using specific FISH probes
to
identify any cytogenetic alterations. The FISH probes are available from
commercial
sources and are stored and used according to the manufacturer's instructions
(either
dual color/dual fusion or dual or tricolor probes):
(i) ATM-p53 DNA probe (ASRs) [del (17); Kreatech, Inc. (Catalog Number
11Q0011495; 17P0011550)
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CA 02915105 2015-12-10
(ii) 4;14 DNA probe (ASRs) [t(4:14)] Kreatech, Inc. (Catalog Number
04P0011495,
14Q0011550)
(iii) 14;16 DNA probe (ASRs)[t(14:16)] Kreatech, Inc. (Catalog Number
16Q001149,
14Q0011550)
[00110] For each probe typically a total of 100 nuclei are counted by 2
technologists using
fluorescent microscopes. If their scores disagree a third technologist counts
additional nuclei.
The following cut-offs are used to score a slide as abnormal (positive for the
cytogenetic
alteration):
(i) ATM/p53 del (17) >5
(ii) 4;14 DNA probe [t(4:14)] >3
(iii) 14;16 DNA probe [t(14:16)] >3
Results (study C16010)
1001111 At the first interim analysis 722 patients had been randomized; 360 in
the
ixazomib arm and 362 in the placebo comparator arm. Based primarily on central
lab
evaluation (97%), 19% of patients were determined to have high-risk
cytogenetics by FISH
[del (17), t(4:14) or t(14;16) including 10% del (17)].
1001121 Analysis of patients defined by at least del (17) showed an
improvement in median
PFS for patients on the ixazomib arm versus the placebo arm (21.4 months vs
9.7 months, HR
= 0.596). Analysis of patients defined by any of the three high-risk
cytogenetic alterations
(del (17), t(4:14) and t(14:16) showed an improvement in PFS for the ixazomib
arm versus
placebo arm (21.4 months vs 9.7 months, HR = 0.543). Additional data for
response rates are
shown below in Table 2
Table 2:
ORR, % >VGPR, % ?CR, % Median PFS, months
IRd Placeb IRd Placeb IRd Placeb IRd Placeb HR
o-Rd o-Rd o-Rd o-Rd
All high- 79* 60 45* 21 12* 2 21.4 9.7 0.54
risk patients 3
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CA 02915105 2015-12-10
Patients 72 48 39 15 11* 0 21.4 9.7 0.59
with 6
del(17p)t
Patients 89 76 53 28 14 4 18.5 12.0 0.64
with t(4;14) 5
alone
*p<0.05 for comparison between regimens. Alone or in combination with t(4;14
or t(14;16);
ORR = overall response rate, VGPR = very good partial response, CR = complete
response.
1001131 As described above a cut-off value of 5 cells per hundred (5%) was
used to
classify a patient with a del (17). Further analysis of the data using cut-
offs of 20% and 60%
is shown in Table 3 below.
Table 3:
Cut-off for del(17p) Number Median PFS, mos HR p-value
of IRd vs placebo-Rd
patients
5% 69 21.4 vs 9.7 0.596 0.162
20% 59 21.4 vs 6.7 0.611 0.2049
60% 33 15.7 vs 5.1 0.49 0.2481
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