U.S. patent application number 10/302654 was filed with the patent office on 2003-06-26 for vitamin d and its analogs in the treatment of tumors and other hyperproliferative disorders.
This patent application is currently assigned to Oregon Health & Science University. Invention is credited to Beer, Tomasz M., Henner, William D..
Application Number | 20030119795 10/302654 |
Document ID | / |
Family ID | 22152202 |
Filed Date | 2003-06-26 |
United States Patent
Application |
20030119795 |
Kind Code |
A1 |
Henner, William D. ; et
al. |
June 26, 2003 |
Vitamin D and its analogs in the treatment of tumors and other
hyperproliferative disorders
Abstract
Treatment of hyperproliferative disorders (including tumors and
psoriasis) by pulse administration of a drug (such as Vitamin D or
an analog) that increases blood or tissue levels of Vitamin D. The
drug is administered at a sufficient dose to have an
anti-proliferative effect, but the pulsed administration of the
drug avoids the development of severe symptomatic or
life-threatening hypercalcemia. In particular embodiments,
avoidance of hypercalcemia (as measured by serum levels of calcium
above normal range) is avoided altogether. In a particular example,
the drug is calcitriol administered at an oral dose of about 0.5
mcg/kg once a week.
Inventors: |
Henner, William D.;
(Portland, OR) ; Beer, Tomasz M.; (Portland,
OR) |
Correspondence
Address: |
Klarquist Sparkman, LLP
One World Trade Center
Suite 1600
121 S.W. Salmon Street
Portland
OR
97204
US
|
Assignee: |
Oregon Health & Science
University
|
Family ID: |
22152202 |
Appl. No.: |
10/302654 |
Filed: |
November 21, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10302654 |
Nov 21, 2002 |
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09647075 |
Sep 21, 2000 |
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6521608 |
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09647075 |
Sep 21, 2000 |
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PCT/US99/06442 |
Mar 25, 1999 |
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60079696 |
Mar 27, 1998 |
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Current U.S.
Class: |
514/167 |
Current CPC
Class: |
A61P 3/14 20180101; A61K
31/59 20130101; A61P 35/00 20180101; A61P 17/12 20180101; A61P
35/02 20180101; A61K 31/593 20130101; A61P 17/06 20180101 |
Class at
Publication: |
514/167 |
International
Class: |
A61K 031/59 |
Claims
We claim:
1. A method for the treatment of a hyperproliferative disease in a
subject, wherein the hyperproliferative disease responds to
treatment with a Vitamin D drug, comprising administering to the
subject a therapeutically effective pulse dose of the Vitamin D
drug in a sufficient amount to have an antiproliferative effect,
without inducing severe symptomatic hypercalcemia.
2. The method of claim 1, comprising administering the Vitamin D
drug to a subject having a neoplasm that expresses a Vitamin D
receptor.
3. The method of claim 2, wherein the neoplasm is selected from the
group of cancer of the prostate, breast, colon, lung, head and
neck, pancreas, endometrium, bladder, cervix, ovaries, squamous
cell carcinoma, renal cell carcinoma, myeloid and lymphocytic
leukemia, lymphoma, medullary thryoid carcinoma, melanoma, multiple
myeloma, retinoblastoma, and sarcomas of the soft tissues and
bone.
4. The method of claim 3, wherein the neoplasm is breast cancer or
prostate cancer.
5. The method of claim 1, wherein the Vitamin D drug has a calcemic
index greater than or equal to calcipitriol.
6. The method of claim 1, wherein the Vitamin D drug has a calcemic
index greater than 1.0.
7. The method of claim 5, wherein the Vitamin D drug has a
half-life that is no greater than about 1 day.
8. The method of claim 7, wherein the Vitamin D drug has a
half-life that is no greater than about 6 hours.
9. The method of claim 8, wherein the Vitamin D drug is
administered in an amount that raises a serum level of Vitamin D in
the subject with a tumor to a supraphysiologic amount for a
sufficient period of time to induce differentiation or regression
of the tumor without causing symptomatic hypercalcemia.
10. The method of claim 1, wherein the Vitamin D drug is
calcitriol, which is administered in a therapeutically effective
pulse dose no more than once every three days.
11. The method of claim 10, wherein the calcitriol is administered
orally in a dose of at least 0.12 mcg/kg per day no more than once
per week.
12. The method of claim 10, wherein the calcitriol is administered
orally in a dose of at least 0.48 mcg/kg or about 1 mcg/kg per day
no more than once per week.
13. A method of treating a tumor in a subject, wherein the tumor
expresses a Vitamin D receptor and is responsive to treatment with
a Vitamin D drug, the method comprising administering orally to the
subject, no more than once every three days, a dose of calcitriol
of about 0.5 mcg/kg.
14. The method of claim 13, wherein the Vitamin D drug is
administered to the subject no more than once per week.
15. A composition comprising a Vitamin D drug in a pharmaceutical
dosage form in a dosage that would cause hypercalcemia is
administered daily.
16. The composition of claim 15, wherein the Vitamin D drug is
calcitriol, contained in the composition in an amount of at least 5
mcg.
17. The composition of claim 16, wherein the pharmaceutical dosage
form is an oral dosage form containing at least 100 mcg per unit of
the oral dosage form.
18. The composition of claim 16, wherein the pharmaceutical dosage
form is a tablet or capsule.
19. The composition of claim 16, wherein the Vitamin D drug is
calcitriol, and the pharmaceutical dosage form is a tablet
containing at least 5 mcg calcitriol.
20. The method of claim 1, wherein the subject is prescribed a
reduced calcium diet for a sufficient period of time prior to
administration of the Vitamin D drug to reduce absorption of
dietary calcium.
21. A method of treating in a subject a tumor that expresses a
Vitamin D receptor, the method comprising raising a blood level of
Vitamin D to a sufficiently supraphysiologic level for a sufficient
period of time to inhibit growth of the tumor, without inducing
hypercalcemia in the subject.
22. The method of claim 21, wherein the blood level of Vitamin D is
raised by administering a Vitamin D drug to the subject.
23. The method of claim 22, wherein the Vitamin D drug is
calcitriol.
24. The method of claim 23, wherein the calcitriol is administered
in a dose of about 0.50 mcg/kg once per week.
Description
FIELD OF THE INVENTION
[0001] This invention concerns the use of Vitamin D and its analogs
in the treatment of tumors and hyperproliferative disorders.
BACKGROUND OF THE INVENTION
[0002] Vitamin D is a generic term for a family of secosteroids
that have affinity for the Vitamin D receptor, and are involved in
the physiologic regulation of calcium and phosphate metabolism.
Exposure to the sun and dietary intake are common sources of
Vitamin D, but deficiencies of this vitamin can cause rickets and
osteomalacia. Supplementation of dairy and other food products has
reduced the incidence of Vitamin D deficiency conditions in modem
society, and medical research concerning this vitamin has turned to
its therapeutic effects in a variety of pathological
conditions.
[0003] Vitamin D.sub.3 is synthesized in human skin from
7-dehydrocholesterol and ultraviolet light. Vitamin D.sub.3, or its
analog Vitamin D.sub.2, can be ingested from the diet, for example
in fortified milk products. Vitamin D.sub.2 and D.sub.3 undergo
hydroxylation first in the liver to 25-hydroxyvitamin D, then in
the kidney to 1.alpha.,25-dihydroxycholecalciferol (also known as
1,25-dihydroxyvitamin D or calcitriol), which is the principal
biologically active form of Vitamin D. The biological production of
this active form of the vitamin is tightly physiologically
regulated.
[0004] Vitamin D exerts its calcium regulating activity through
both genomic and nongenomic pathways. Although the nongenomic
pathways remain poorly characterized, the genomic responses are
mediated through binding to the nuclear Vitamin D receptor (VDR).
The VDR is a ligand-activated transcription factor, which binds the
Vitamin D.sub.3 response element contained within the
promoter/enhancer region of target genes. Vitamin D maintains
calcium levels in the normal range by stimulating intestinal
calcium absorption. When intestinal absorption is unable to
maintain calcium homeostasis, Vitamin D stimulates monocytic cells
to become mature osteoclasts, which in turn mobilize calcium from
bones.
[0005] Appreciation for Vitamin D's non calcium-related biological
activities began in 1979 with Stumpf's discovery that radioactive
Vitamin D localizes to many tissues not associated with calcium
metabolism (Science 206:1188-1190, 1979). In 1981, Abe et al.
reported that mouse myeloid leukemia cells possessed VDR, and that
their exposure to Vitamin D led to terminal differentiation (PNAS
USA 78:4990-4994, 1981). Since then VDR has been described in
carcinomas of the prostate, breast, colon, lung, pancreas,
endometrium, bladder, cervix, ovaries, squamous cell carcinoma,
renal cell carcinoma, myeloid and lymphocytic leukemia, medullary
thyroid carcinoma, melanoma, multiple myeloma, retinoblastoma, and
sarcomas of the soft tissues and bone.
[0006] In vitro assays using 1,25 dihydroxyvitamin D or its
analogues demonstrated antiproliferative effects in cell lines
derived from many malignancies including adenocarcinomas of the
prostate (Molec. and Cell. Endocrinology 126:83-90, 1997; Proc.
Amer. Assoc. Cancer Res. 38:456, 1997; J. Ster. Biochem. and Molec.
Biol. 58:277-288, 1996; Endocrinology 137:1554-1561, 1996;
Endocrinology 136:20-26, 1995; Cancer Research 54:805-810, 1994;
Endocrinology 132:1952-1960, 1993; and Anticancer Research
14:1077-1081, 1994), breast (Proc. Amer. Assoc. Cancer Res. 38:456,
1997; Biochemical Pharmacology 44:693-702, 1992); colon
(Biochemical and Biophysical Research Communications 179:57-62,
1991; Archives of Pharmacology 347:105-110, 1993); pancreas
(British Journal of Cancer 73:1341-1346, 1996); and endometrium
(Journal of Obstetrics and Gynaecology Research 22:529-539, 1996);
lung (Anticancer Research 16:2953-2659, 1996); myeloid leukemia
(PNAS USA 78:4990-4994, 1981); melanoma (Endocrinology
108:1083-1086, 1981); and sarcomas of the soft tissues (Annals of
Surgical Oncology 3:144-149, 1996) and bone (Journal of the
Japanese Orthopaedic Association 69:181-190, 1995).
[0007] Studies in animals have shown antiproliferative activity of
Vitamin D or its analogues in prostate cancer (Urology 46:365-369,
1994); breast cancer (J. NCI 89:212-218, 1997; Lancet 1: 188-191,
1989); squamous cell carcinoma (Molecular and Cellular
Differentiation 3:31-50, 1995); myeloid leukemia (Blood 74:82-93,
1989 and PNAS USA 80:201-204, 1983) and retinoblastoma (Archives of
Opthalmology 106:541-543, 1988; Archives of Opthalmology
106:536-540, 1988). The mechanism of Vitamin D's antiproliferative
effects remains unknown, although it has been proposed that Vitamin
D increases synthesis of TGF-.beta.1 and TGF-.beta.2, decreases the
expression of epidermal growth factor receptors, leads to
dephosphorylation of the retinoblastoma protein, induces cell cycle
arrest in G1, perhaps by induction of the cyclin dependent kinase
inhibitors p21 (waf1) and p27 (kip1), and induces the production of
insulin-like growth factor binding protein.
[0008] The patent literature is replete with attempts to treat
tumors with Vitamin D compounds. U.S. Pat. No. 4,391,802 disclosed
treating leukemioid diseases with 1.alpha.-hydroxy Vitamin D
derivatives. The use of 1.alpha.-hydroxy derivatives with a 17 side
chain greater in length than the cholesterol or ergosterol side
chains was disclosed in U.S. Pat. No. 4,717,721. Additional Vitamin
D analogs are described in U.S. Pat. No. 4,851,401
(cyclopentano-Vitamin D analogs), U.S. Pat. No. 4,866,048, U.S.
Pat. No. 5,145,846 (Vitamin D.sub.3 analogs with alkynyl, alkenyl,
and alkanyl side chains), U.S. Pat. No. 5,120,722
(trihydroxycalciferol), U.S. Pat. No. 5,547,947
(fluorocholecalciferol compounds), U.S. Pat. No. 5,446,035 (methyl
substituted Vitamin D), U.S. Pat. No. 5,411,949
(23-oxa-derivatives), U.S. Pat. No. 5,237,110 (19-Nor-Vitamin D
compounds), U.S. Pat. No. 4,857,518 (hydroxylated 24-homo-Vitamin D
derivatives). Additional Vitamin D analogs are shown in U.S. Pat.
Nos. 4,804,502; 5,374,629; 5,403,940; 5,446,034; and 5,447,924.
[0009] Few attempts have been made to test Vitamin D's
antiproliferative effects in humans with cancer. Koeffler et al.,
Cancer Treatment Reports 69:1399-1407, 1985, gave 2 mcg of
1,25-dihydroxyvitamin D daily for 8 weeks or longer to 18 patients
with myelodysplastic syndrome. Eight of 18 patients had minor and
transient improvements in the peripheral blood counts, but by the
end of the 12 week study no patient showed significant improvement
and 4 patients experienced symptomatic hypercalcemia. Bower et al.,
Lancet 337:701-702, 1991, treated 19 patients with locally advanced
or cutaneous metastatic breast cancer with topical calcipotriol, a
Vitamin D analogue. Three of the 14 patients who completed 6 weeks
of treatment showed a 50% reduction in the bidirectional diameter
of the treated lesions and one other patient showed minimal
response, however hypercalcemia was a complication of the
treatment. Palmieri-Sevier et al., Am. J. Medical Sciences
306:309-312, 1993, reported a case of long term remission of
parathyroid carcinoma which appeared to be induced and maintained
by Vitamin D therapy. Rustin et al., Brit. J. Can. 74:1479-1481,
1996, performed a clinical trial with a continuous dose of
calcitriol in patients with ovarian cancer, and again encountered
hypercalcemia.
[0010] A phase II trial of oral 1,25-dihydroxyvitamin D
(calcitriol) in hormone refractory prostate cancer was reported by
Osborn et al., Urol. Oncol., 1: 195-198, 1995. Fourteen patients
were given a daily oral dose of 0.5-1.5 mcg calcitriol, but no
significant response was demonstrated, and clinical deterioration
was documented in most of the patients. Thirteen of the patients
experienced hypercalcemia, which is the most common side effect of
treatment with Vitamin D and its analogs. Concern that
hypercalcemic effects of Vitamin D would preclude the achievement
of therapeutic, anti-neoplastic serum levels has inhibited the
study of the use of this vitamin in humans with cancer. It is an
object of this invention to provide a method of treatment with
Vitamin D drugs (such as calcitriol) that avoids such
hypercalcemia, while permitting the use of this class of drugs in
the treatment of tumors and other hyperproliferative diseases.
SUMMARY OF THE INVENTION
[0011] Vitamin D and its analogs can be administered in accordance
with the present invention, for the treatment of neoplastic
diseases, such as the types of tumors mentioned above, which are
responsive to treatment with Vitamin D drugs. The method can also
be used to treat hyperproliferative skin diseases, such as
psoriasis, disorders of keratinization and keratosis, or disorders
of sebaceous glands, such as acne or seborrheic dermatitis. The
method includes administering to the subject a therapeutically
effective pulsed dose of the Vitamin D drug in a sufficient amount
to have a therapeutic effect, without inducing hypercalcemia,
particularly symptomatic hypercalcemia, for example stage 3 or
stage 4 hypercalcemia. This treatment is especially effective to
allow the use of highly calcemic drugs (such as drugs having a
calcemic index of 0.5 or more) which are often highly effective in
the treatment of tumors and hyperproliferative diseases, but which
have been avoided in the past because of their calcemic side
effects. The dosing regimen of the present invention for the first
time allows therapeutically effective antiproliferative (and
particularly antineoplastic) amounts of these drugs to be given
without inducing the dangerous side effect of life-threatening
hypercalcemia, while surprisingly having a prolonged therapeutic
specific anti-tumor or general antiproliferative effect.
[0012] In a first disclosed embodiment, the Vitamin D drug is
administered to a subject having a neoplasm that expresses a
Vitamin D receptor, and responds to treatment with a Vitamin D
drug. Particular types of tumor that respond to such treatment
include adenocarcinomas of the prostate, breast, colon, pancreas
and endometrium, as well as small cell and non-small cell cancer of
the lung (including squamous, adenocarcinoma and large cell types),
squamous cell carcinoma of the head and neck, transitional cell
cancer of the bladder, ovarian and cervical (e.g. squamous cell
carcinoma) cancer, renal cell carcinoma, myeloid and lymphocytic
leukemia, lymphoma, medullary thyroid carcinoma, melanoma, multiple
myeloma, retinoblastoma, and sarcomas of the soft tissues and bone.
In particular embodiments, the neoplasm is adenocarcinoma of the
breast or prostate.
[0013] In yet other specific embodiments, the Vitamin D drug is one
that would induce hypercalcemia (particularly symptomatic or
life-threatening hypercalcemia) in a subject to which the drug is
given at antiproliferative doses. The method would have particular
application to drugs that are as calcemic as calcipotriol (calcemic
index of about 0.005-0.01),
11.alpha.-fluoromethyl-1.alpha.,25-(OH).sub.2-D.sub.3 (having a
calcemic index of about 0.1), and drugs having a calcemic index
greater than 0.5, for example greater than or equal to 1 (the
calcemic index of calcitriol). Drugs with which the method is
particularly useful are those drugs having a half-life no greater
than about 1 day, for example no greater than about 6 hours, when
the dose is given as a therapeutically effective dose. These
half-lives are sufficiently short that they allow the blood level
to return to non-calcemic levels for a sufficient period between
doses so that full osteoclast activation does not occur. In
particular embodiments, blood levels of calcium return to normal
between doses. The Vitamin D drug is administered in an amount that
raises a serum level of Vitamin D in the subject with a tumor to a
supraphysiologic amount for a sufficient period of time to induce
differentiation or regression of the tumor without causing
symptomatic hypercalcemia.
[0014] For example, where the Vitamin D analog is calcitriol, it
can be administered in a high pulse dose no more than once every
three days, for example once a week. Although calcitriol has been
used in the past to treat cancer, dosages of such regimens have
been 0.5-1.5 mcg per day for prolonged periods of time, which has
caused symptomatic hypercalcemia. In accordance with some
embodiments of the present invention, calcitriol is orally
administered in a dose of at least 0.12 mcg/kg per day (8.4 mcg in
a 70 kg person) no more than once every 5 or 6 days, for example
once a week. Even higher doses of calcitriol are possible using the
pulsed dose regimen of the present invention, for example
administering the calcitriol orally in a dose of about at least
about 0.48 mcg/kg per day, for example 1 mg/kg per day or higher,
e.g. 2-3 mg/kg per day, no more than once per week. As the dosage
of the calcitriol or other Vitamin D drug increases, the interval
between doses can be increased (for example to as long as 7-10
days) to avoid symptomatic hypercalcemia. It has surprisingly been
observed that pulsed increases in the blood level of Vitamin D are
sufficient to have an anti-tumor or antiproliferative effect for a
prolonged period of time (e.g. 10 days), so that the dosing regimen
of the present invention can be followed while encountering a
lowered risk of hypercalcemia.
[0015] The invention also includes a pharmaceutical composition
comprising a Vitamin D drug in a pharmaceutical dosage form
containing at least 5 micrograms (mcg) of calcitriol, for example
5-100 mcg. The dosage form may be an oral, intravenous,
intramuscular, topical, subcutaneous, transdermal, sublingual,
intranasal, intratumoral or other preparation, but in particular
disclosed embodiments the pharmaceutical dosage form is an oral
dosage form, such as a tablet or capsule.
[0016] The foregoing and other objects, features, and advantages of
the invention will become more apparent from the following detailed
description of several preferred embodiments.
BRIEF DESCRIPTION OF THE FIGURES
[0017] FIG. 1 is a diagram showing peak and trough plasma
calcitriol levels in subjects who received the indicated dose of
calcitriol over four hours. Peak levels (.diamond.) were determined
at 6 hours after administration, and trough levels (.largecircle.)
were determined at 48 hours after administration.
[0018] FIG. 2 shows a time course of plasma calcitriol levels in a
subject who received a 2.0 .mu.g/kg dose of calcitriol.
DETAILED DESCRIPTION OF SEVERAL EMBODIMENTS
[0019] Definitions The following definitions will help with an
understanding of the terms used in this specification.
[0020] A "Vitamin D drug" is a drug that raises the blood or tissue
level of Vitamin D, or has an affinity for the Vitamin D receptor,
for example binding to that receptor with a Relative Competitive
Index (RCI) of 0.05 or greater, more particularly 5 or greater, for
example 5-250. The RCI is indexed to an RCI of 100 for calcitriol.
The term also includes any of the family of secosteroids with
antirhichitic activity, such as Vitamin D.sub.2 (ergocalciferol)
and Vitamin D.sub.3 (cholecalciferol), their precursor molecules
such as ergosterol (7-dehydro-22-dehydro-24-methyl-ch- olesterol)
and 7 dehydrocholesterol, 25-hydroxyvitamin D.sub.3 the
3-hydroxylated dihydrotachysterol.sub.2, the 1.alpha.-hydroxylated
alfacalcidol (1.alpha.-hydroxyvitamin D.sub.3) and calcitriol
(1.alpha.,25-dihydroxyvitamin D.sub.3), as well as the numerous
natural and synthetic Vitamin D analogs set forth in the attached
Appendix I (from Bouillon et. al, Endocrine Reviews 16:
200-257,1995).
[0021] Vitamin D drugs also include Vitamin D preparations and
analogs that are currently in clinical use, such as Rocaltrol.RTM.
(Roche Laboratories), Calcijex.RTM. injectable calcitriol,
investigational drugs from Leo Pharmaceutical including EB 1089
(24a,26a,27a-trihomo-22,24-dien-
e-1.alpha.a,25-(OH).sub.2-D.sub.3), KH 1060
(20-epi-22-oxa-24a,26a,27a-tri-
homo-1.alpha.,25-(OH).sub.2-D.sub.3), MC 1288 and MC 903
(calcipotriol), Roche Pharmaceutical drugs that include
1,25-(OH).sub.2-16-ene-D.sub.3,
1,25-(OH).sub.2-16-ene-23-yne-D.sub.3, and
25-(OH)2-16-ene-23-yne-D.sub.3- , Chugai Pharmaceuticals
22-oxacalcitriol (22-oxa-1.alpha.,25-(OH)2-D.sub.- 3;
1.alpha.-(OH)D.sub.5 from the University of Illinois; and drugs
from the Institute of Medical Chemistry-Schering AG that include ZK
161422 and ZK 157202. Appendix 3 provides additional information
about chemical structure, route of administration and dosing of
some of these compounds. Vitamin D analogs also include topical
preparations of Vitamin D, such as Calcipotriene (Dovonex) and
Tacalcitol, used in the treatment of psoriasis.
[0022] A "Vitamin D receptor" (or VDR) is a protein transcription
factor, for which the gene and its product have already been
characterized and found to contain 427 amino acids, with a
molecular weight of about 47,000, or variants thereof The full
length cDNA of the human VDR is disclosed in Baker et al., PNAS,
USA 85:3294-3298, 1988.
[0023] "Tumor cells that express (or contain) the Vitamin D
receptor" are those tumors that have been shown to contain the
Vitamin D receptor, tumors that are subsequently shown to contain
the receptor (using immunohistochemical or other techniques), tumor
types (such as breast cancer) that have demonstrated a clinical
improvement in response to treatment with calcitriol or its analogs
or other Vitamin D drugs, and tumors for which there is
epidemiologic data demonstrating an association between low Vitamin
D levels and higher cancer incidence (such as adenocarcinomas of
the prostate, breast and colorectum). The presence of Vitamin D
receptors can be determined by any means known in the art, such as
any of the techniques disclosed in Pike, Ann Rev. Nut. 11: 189-216,
1991.
[0024] "Elevated (or supraphysiologic) level of Vitamin D" refers
to a 1,25-dihydroxyvitamin D plasma concentration greater than
about 0.15 nm (65 pg/ml), or other Vitamin D concentration greater
than normal in the laboratory where the concentration is measured,
for example in humans a total human plasma concentration greater
than about 10 ng/ml of 25-hydroxyvitamin D (although this and all
other normal values can vary depending on the techniques used to
measure the concentration).
[0025] "Hypercalcemia" refers to a calcium plasma concentration
greater than normal in the laboratory where the concentration is
measured, for example greater than about 10.5 mg/dL in humans
(although this and all other normal values can vary depending on
the techniques used to measure the concentration). Hypercalcemia
can be broken into grades 0-4, as set forth in Appendix II.
[0026] "Symptomatic hypercalcemia" refers to laboratory
demonstrated hypercalcemia associated with one of more of the signs
or symptoms of hypercalcemia. Early manifestations of hypercalcemia
include weakness, headache, somnolence, nausea, vomiting, dry
mouth, constipation, muscle pain, bone pain, or metallic taste.
Late manifestations include polydypsia, polyuria, weight loss,
pancreatitis, photophobia, pruritis, renal dysfunction,
aminotransferase elevation, hypertension, cardiac arrhythmias,
psychosis, stupor, or coma. Ectopic calcification has been reported
when the calcium-phosphate product (multiplying the concentrations
of calcium and phosphate) exceeds 70. "Severe symptomatic
hypercalcemia" refers to grade 3 or grade 4 hypercalcemia.
[0027] A "tumor" is a neoplasm, and includes both solid and
non-solid tumors (such as hematologic malignancies). A
"hyperproliferative disease" is a disorder characterized by
abnormal proliferation of cells, and generically includes skin
disorders such as psoriasis as well as benign and malignant tumors
of all organ systems. "Differentiation" refers to the process by
which cells become more specialized to perform biological
functions, and differentiation is a property that is totally or
partially lost by cells that have undergone malignant
transformation.
[0028] A "therapeutically effective dose" is a dose which in
susceptible subjects is sufficient to prevent advancement, or to
cause regression of the disease, or which is capable of relieving
symptoms caused by the disease, such as fever, pain, decreased
appetite or chachexia associated with malignancy.
[0029] A "pulse" dose of a Vitamin D drug refers to administration
of the drug in a sufficient amount to increase the blood or tissue
level of Vitamin D to a supraphysiologic concentration for a
sufficient period of time to have a therapeutic benefit, but with a
sufficient period between doses to avoid hypercalcemia, given the
pharmacological half life of the drug, its rate of elimination from
the body, and its calcemic index.
[0030] The "calcemic index" of a drug is a measure of the relative
ability of a drug to generate a calcemic response, for example as
measured and reported in Bouillon et al., Endocrine Reviews
16:200-257, 1995. A calcemic index of 1 corresponds to the relative
calcemic activity of calcitriol. A calcemic index of about 0.01
corresponds to the calcemic activity of calcipotriol. A calcemic
index of 0.5 would correspond to a drug having approximately half
the calcemic activity of calcitriol. The calcemic index of a drug
can vary depending on the assay conducted, e.g. whether measuring
stimulation of intestinal calcium absorption (ICA) or bone calcium
mobilizing activity (BCM), as reported in Hurwitz et al., J. Nutr
91:319-323, 1967 and Yamada et al., Molecular, Cellular and
Clinical Endocrinology (Berlin), pages 767-774, 1988. Hence
relative calcemic activity is best expressed in relation to the
calcemic activity of calcitriol, which is one of the best
characterized Vitamin D drugs.
Vitamin D Drugs
[0031] Normal serum levels of 1,25-dihydroxyvitamin D range between
0.05 and 0.16 nM, however therapeutic drug levels necessary for
cancer inhibition have not been well defined. Skowronski et al.
(Endocrinology 136-20-26, 1995) demonstrated measurable growth
inhibition of LNCaP human prostate cancer cells in vitro at a
1,25-dihydroxyvitamin D concentration of 0.1 nM and 50% growth
inhibition at a 1.0 nM concentration. Peehl et al. (Cancer Research
54:805-810, 1994) incubated human prostate cancer cells in primary
culture with 1,25-dihydroxyvitamin D concentrations ranging between
0.025 and 25 nM and demonstrated half maximal growth inhibition at
levels between 0.25 and 1.0 nM. Previous clinical trials of Vitamin
D in the treatment of cancer have proceeded on the assumption that
high levels of the drug were needed for a prolonged period of time
to have a therapeutic benefit. The inventors of the present
invention, however, have surprisingly shown that intermittent
supraphysiologic levels of 1,25-dihydroxyvitamin D (for example
greater than or equal to 0.25 nM) are sufficient to inhibit cancer
growth and other proliferative disease in mammals. This surprising
finding now permits the therapeutic benefits of Vitamin D therapy
to be achieved without substantial risk of morbidity from
iatrogenic hypercalcemia induced by the therapy.
[0032] Calcitriol is a short acting preparation of
1,25-dihydroxyvitamin D, which therefore offers an opportunity for
intermittent treatment aimed at achieving high serum
1,25-dihydroxyvitamin D levels for brief periods of time. This
regimen has surprising anti-tumor activity, while minimizing
toxicity, such as hypercalcemia. Calcitriol has primarily been
studied when chronically administered as replacement therapy, for
which its usual dose is 0.25-1.0 mcg per day. Peak serum
concentration is reached at 2 hours and serum half life is 3-6
hours. Intestinal absorption of calcium begins to increase 2 hours
after administration. Hypercalcemic effect is maximal at 10 hours
and lasts 3-5 days.
[0033] In one embodiment of the invention, a sufficient dose of
calcitriol is administered to raise serum 1,25-hydroxyvitamin D
levels to a therapeutically effective level for a pulsed dose that
has an anti-proliferative effect without causing significant
hypercalcemia (for example symptomatic grade 3 or grade 4
hypercalcemia). With calcitriol, an example of such a dose would
produce a serum level of at least about 0.5 nM, for example about
0.9 nM or more (e.g. 1-25 nM, for example 5-10 nM), for at least 2
hours (e.g. 2-5 hours) and preferably no more than 6 hours. In
particular embodiments, the pulsed dose of calcitriol does not
exceed a dose at which symptomatic hypercalcemia occurs, or more
preferably a pulsed dose at which even laboratory hypercalcemia
occurs.
[0034] Information about short term effects of higher than
replacement doses of calcitriol is available for helping predict
drug effects. Papapoulus et al., (Clinical Science 62:427-429,
1982) gave 2 mcg of calcitriol as a single oral dose to healthy
volunteers and achieved peak 1,25-dihydroxyvitamin D serum
concentrations of 0.235 and 0.351 nM. Mason et al. (BMJ
1980:449-450) gave a single oral dose of 4 mcg calcitriol to
healthy volunteers and achieved peak 1,25-dihydroxyvitamin D serum
concentrations of 0.42 nM with no elevation in serum calcium.
Brickman et al. (Am J. Med. 57:28-33, 1974) treated normal
volunteers with calcitriol doses up to 2.7 mcg/day for 7 to 15
days. While calcium absorption and excretion were increased, no
significant elevations in serum calcium were observed. Adams et al.
(Kidney Int. 21:90-97, 1982) treated normal volunteers with up to 3
mcg/day of calcitriol for 6-12 days and achieved stable
1,25-dihydroxyvitamin D serum levels of 0.184-0.235 nM. None of the
patients who were on a low calcium diet experienced elevation in
serum calcium. Geusens et al. (Calcified Tissue Int. 49:168-173,
1991) gave 4 mcg of calcitriol per day for 4 days to 27
postmenapausal women with osteoporosis or osteoarthritis. They
demonstrated increased urinary calcium excretion but no increase in
urinary hydroxyproline excretion. Four of the 27 patients had a
serum calcium above 10.8 but no clinically significant
hypercalcemia was observed.
[0035] Antiproliferative levels of 1,25-dihydroxyvitamin D can be
achieved for short periods of time with minimal adverse effects,
particularly if hypercalcemia during short course
1,25-dihydroxyvitamin D therapy is primarily mediated by increases
in intestinal calcium absorption (slower calcium elevation) rather
then osteoclast activation (which can rapidly mobilize calcium from
bone). Higher 1,25-dihydroxyvitamin D levels are achievable when
the drug is given in conjunction with a reduced calcium diet to
minimize intestinal calcium absorption, and adequate hydration to
maximize calcium excretion. The maximal tolerated dose of
calcitriol, when given intermittently has not been defined, but
doses as high as 0.48 mcg/kg have been tolerated without
hypercalcemia.
[0036] Higher doses of a Vitamin D drug, sufficient to achieve
therapeutic antiproliferative levels, may also be achieved by
administering the drug in conjunction with bisphosphonate
osteoclast inhibitors, such as pamidronate. Selby et al.
(Endocrinology 108:1083-1086, 1981) provided an example of treating
hypercalcemia due to Vitamin D with pamidronate. The potential for
achieving high serum 1,25-dihydroxyvitamin D levels when
osteoclasts are inhibited in patients with osteopetrosis is
possible with calcitriol doses as high as 32 mcg/day for 3 months
(Key et al., NEJM 310:409-415, 1984) where stable serum levels of
1,25-dihydroxyvitamin D peaked at 2.32 nM with no
hypercalcemia.
[0037] The following Examples illustrate the general method of the
present invention, as well as specific case histories to illustrate
its use. These Examples also provide a general framework for
evaluating other Vitamin D drugs, and determining a therapeutically
effective dose of a Vitamin D drug in a subject with a Vitamin D
responsive hyperproliferative disease, without inducing symptomatic
iatrogenic hypercalcemia.
EXAMPLE 1
General Treatment Plan
[0038] A patient with a known Vitamin D receptor positive tumor
(such as adenocarcinoma of the prostate, breast, lung, colon or
pancreas, or transitional cell carcinoma of the bladder, or
melanoma) may be placed on a prescribed reduced calcium diet prior
to treatment, to help minimize intestinal absorption and allow even
higher doses of the Vitamin D drug to be used. This reduced calcium
diet may be continued for the duration of treatment, and for one
week after the last dose of the Vitamin D drug. The diet ideally
restricts daily calcium intake to 400-500 mg, by avoiding all dairy
products, as well as sardines and other fish canned with their
bones, legumes, greens, and any calcium fortified foods or drinks.
The subject is then asked to drink 4-6 cups of fluid more than
usual intake starting 12 hours before treatment and continuing for
days 1, 2, and 3, to assure adequate oral hydration. Magnesium
containing antacids, oral calcium supplements, cholestyramine,
colestipol, and other bile resin binding agents may also be avoided
during treatment.
[0039] Baseline laboratory tests that may be obtained include serum
levels of calcium, phosphate, and 1,25-dihydroxyvitamin D. At the
initial dose level, e.g. calcitriol 0.06 mcg/kg po (or another
Vitamin D drug for which the dose is to be determined) is divided
into 4 doses, and one of those four doses is taken during each hour
for 4 hours until the total 0.06 mcg/kg dose is taken.
Alternatively, a single higher dose formulation may be ingested.
The doses may be rounded up to the nearest 0.5 mcg. The subject is
monitored daily for symptoms of hypercalcemia for at least 2-3 days
following administration.
[0040] The patient may have a variety of laboratory tests performed
to monitor the presence of hypercalcemia, or any physiological
consequences of hypercalcemia. Such tests may include calcium at 0,
24, 48 hours, and baseline levels of creatinine, total billirubin,
ALT, alkaline phosphatase, and a complete blood count. Other
possible laboratory tests include phosphate, 1,25-dihydroxyvitamin
D levels at 0, 6, 24, 48 hours, and 24 hour urine collection for
calcium and hydroxyproline on day 2. Subjects are treated with the
once a week pulse dose of Vitamin D weekly until disease
progression or 4 weeks, whichever comes first, and are followed for
2 months from enrollment. If Grade 3 toxicity is encountered, the
treatment is stopped.
[0041] An initial dose may be chosen from safe doses documented in
the literature, followed by a multistage escalation scheme, such as
the one described by Gordon and Willson (Statistic in Medicine
11:2063-2075, 1992). Patient accrual occurs in stages I, II, and
III. The stages require the accrual of one, three, or six patients
respectively before dose escalation. All patients enrolled at a
dose level complete 4 weeks of treatment before the dose level is
escalated. In stage I, a single patient is entered at each dose
level. Accrual continues in stage I until the first Grade 3
toxicity is encountered. When a Grade 3 toxicity is encountered,
two more patients are accrued at the same dose level and accrual
will continue in stage II. Doses are reduced one level if one Grade
4 or 5 toxicity is encountered in stage I.
[0042] Accrual continues in stage II if no Grade 3 toxicities are
encountered. When one or two Grade 3 or Grade 4 toxicities are
encountered, three additional patients are accrued at the same dose
level and accrual continues in stage III. Doses are reduced one
level if one Grade 5 or three Grade 3 or Grade 4 toxicities are
encountered in stage II. In stage III six patients are enrolled at
each dose level. If only one Grade 3 toxicity is encountered, the
dose will be escalated and the accrual will revert to stage II. If
two or more Grade 3 or greater toxicities occur, no further
escalation will occur. The MTD (maximum tolerated dose) is defined
as that dose at which 1/3 or fewer of the subjects experience grade
3 toxicity. For calcitriol, the initial dose was 0.06 mcg/kg po
over 4 hours. At each successive level, the dose is doubled until
the first grade 3 toxicity is encountered. After that, each dose
increase is 1.33.times.of the preceding level according to a
modified Fibonacci scheme (Diliman and Koziol, Molecular Biotherapy
4:117-121, 1992).
[0043] For calcitriol, the pulse dose was given to each subject
weekly, and the subject was monitored for early signs and symptoms
of hypercalcemia, such as weakness, headache, somnolence, nausea,
vomiting, dry mouth, constipation, muscle pain, bone pain, metallic
taste. The patient was also monitored for any more serious
manifestations, such as polydypsia, polyuria, weight loss,
pancreatitis, photophobia, pruritis, renal dysfunction,
aminotransferase elevation, hypertension, cardiac arrhythmias,
psychosis, stupor, coma, and ectopic calcification. Appropriate
treatment is instituted for any patient who demonstrates
hypercalcemic toxicity, and the calcitriol is stopped until serum
calcium returns to normal.
[0044] The following Table 1 illustrates a protocol that can be
followed with each drug to determine a tolerated pulse dose. A
protocol for determining a therapeutic dose will be described in
Example 2.
1TABLE 1 Example of Protocol for Determining Tolerated Dose WEEKS
5-8 WEEKS 1-4 Follow-Up Premature EVALUATION Day Day Day Week Week
Until Ca nl After Ca nl Evaluation & Procedures Screen 1 2 3 5
7 daily every 2 Informed Consent x Inclusion/Exclusion Criteria x
Physical Exam x x x x Sitting Vital Signs x x x Adverse Effects
Recorded x x x x x x Calcitriol administered x Calcium x x x x x x
x x Phosphate x x x x x 1,25-dihydroxyvitamin D level x xx x x x
Creatinine x x x Total Billirubin x x x ALT x x x Akaline
Phosphatase x x x Albumin x Complete Blood Count x x .beta.-hCG
(select patients) x Urine Collection x x Diet Questionnaire x Tumor
measurements (when x x appropriate)
EXAMPLE 2
Determination of Therapeutically Effective Dose
[0045] Tumor markers, such as PSA, CA 15-3, and others can be used
to assess tumor progression or regression, although the results of
such assays can sometimes be difficult to interpret because
administration of Vitamin D has been shown to increase tumor marker
production while inhibiting cancer cell growth. This effect is
presumably due to the differentiation inducing properties of
Vitamin D.
[0046] Alternative means for determining a therapeutic response can
also be employed, for example direct radiographic measurement of
tumor lesions. A measurable lesion may be considered one that is
bidimensionally measurable, with clearly defined margins on
physical exams, x-ray, or scan. At least one diameter is preferably
greater than 0.5 cm. Bone lesions are not included.
[0047] Evaluable disease includes unidimensionally measurable
lesions, masses with margins not clearly defined, palpable nodal
disease, lesions with both diameters less than 0.5 cm, and bone
disease. Non-evaluable disease includes disease manifested by
pleural effusions, ascites, or disease documented by indirect
evidence only (e.g., by lab values which fall into a category of
not being evaluable). The objective status is recorded at entry
into the trial and during week 7 (where week 1 is the week during
which the first dose of the Vitamin D drug is given). If an organ
has too many measurable lesions to measure at each evaluation, a
specific number (such as three lesions) are selected to be followed
before the patient is entered in the study.
[0048] A complete response (CR) is the complete disappearance of
all measurable and evaluable disease, with no new lesions. If the
subject has effusions, ascites or disease assessable by surgical
restaging (e.g., testicular and extragonadal gem cell cancer), the
disease must be cytologically negative. Patients with markers or
indirect evidence of involvement must have normalization of
abnormal values. All measurable, evaluable and non-evaluable
lesions and sites must be assessed. A partial response (PR) is
found in subjects with at least one measurable lesion with 050%
decrease of perpendicular diameters of all measurable lesions, with
no progression of evaluable disease, and no new lesions. All
measurable and evaluable lesions and sites must be assessed. In
lung cancer, a greater than 50% decrease in estimated area of
evaluable, but non-measurable, tumor mass, as agreed upon by two
independent observers, not to include pleural effusions.
Stabilization is a response that does not qualify as a complete
response, partial response or progression.
EXAMPLE 3
Treatment of Breast Cancer
[0049] In this example, a 42 year old woman with breast carcinoma
metastatic to numerous sites in the skeleton received a dose of 11
mcg of calcitriol (Rocaltrol, Roche) administered as 22 tablets
(0.5 mcg each tablet) divided into four nearly equal doses given in
hour one, two, three and four. The patient received this same
therapy on day 1, 8, 15 and 22, and then was observed on study
until day 56, and tolerated the treatment well. She had no Grade II
or higher toxicities on the NCI toxicity grading scale (Appendix
2). Subjective beneficial effects observed included a reduction in
pain and in analgesia required. Objective benefits included a
progressive decrease in the serum tumor marker CA15-3 from 445
(pre-treatment) to 365 (day 27), 365 (day 48) and 320 (day 70).
Radiologic evaluation of areas of known bony involvement showed
progressive sclerosis of multiple lesions in the pelvis and right
hip, indicating bone healing as a positive response to therapy. No
new lesions or pathologic fractures identified were identified by
day 64.
EXAMPLE 4
Treatment of Melanoma
[0050] In this example a 72 year old man with metastatic malignant
melanoma of the right jaw received a dose of 37 mcg of calcitriol
(Rocaltrol, Roche) administered as 74 tablets (0.5 mcg each tablet)
divided into four nearly equal doses given in hour one, two, three
and four. The patient received this same therapy on day 1, 8, 15
and 22, and observed until at least day 56. The level of calcitriol
in a plasma sample obtained two hours after the last dose of
calcitriol (on week one) was determined using a commercial assay at
Endocrine Sciences, Inc. The level was 1826 pg/ml, compared to the
range for calcitriol levels in normal controls being 21 to 65
pg/ml. In spite of the markedly elevated levels of calcitriol
achieved with this weekly schedule, this patient did not have any
subjective or objective toxicity. Levels of serum calcium and other
chemical and hematological parameters in the blood remained
normal.
EXAMPLE 5
Summary of Trial Results
[0051] Patients
[0052] Eligibility criteria included histologically confirmed
malignancy refractory to standard therapy; age .gtoreq.18 years;
expected survival of >2 months; ECOG performance status
.ltoreq.2; hematocrit .gtoreq.30; serum creatinine .ltoreq.1.2
mg/dL; serum calcium .ltoreq.10.5 mg/dL; serum phosphate
.ltoreq.4.2 mg/dL; ALT .ltoreq.60 IU/L; total serum bilirubin <2
mg/dL. Exclusion criteria included pregnancy, history of
hypercalcemia, kidney stones, heart failure or significant heart
disease including myocardial infarction in the last 3 months, known
cardiac ejection fraction less than 30%, current digoxin therapy,
thiazide diuretic therapy within 7 days, bisphosphonate treatment
within 4 weeks, systemic steroid therapy within 2 weeks, and
unwillingness or inability to stop all magnesium containing
antacids, bile resin binding drugs, or calcium supplements for the
duration of the study.
[0053] Treatment
[0054] Baseline evaluation included a complete history and physical
exam, complete blood count, serum creatinine, serum calcium, serum
phosphate, total serum bilirubin, ALT, alkaline phosphatase,
albumin, serum .beta.-hCG in women of childbearing potential, 24
hour urine collection for calcium, and tumor measurements.
[0055] Patients were asked to maintain a reduced calcium diet for
the four treatment weeks, with a goal of less than 500 mg per day,
as described in Example 1. Calcitriol (Rocaltrol.RTM., Roche
Pharmaceuticals) was given orally once a week for four weeks. Each
weekly dose was given in four divided doses given hourly over four
hours. The starting dose was 0.06 .mu.g/kg.
[0056] Monitoring
[0057] Complete blood count, serum creatinine, total serum
bilirubin, ALT, alkaline phosphatase were monitored weekly. Serum
calcium and phosphate were checked on the treatment day (day 1),
and on days 2 and 3. Twenty-four hour urinary calcium excretion was
measured on day 2. The 1,25-dihydroxyvitamin D levels were measured
by .sup.125I radioimmunoassay (Incstar, Stillwater, Minn.) and by a
radioreceptor assay using calf thymus 1,25-dihydroxyvitamin D
receptor (Endocrine Sciences, Calabasas Hills, Calif.). Peak levels
were measured two hours after all the pills had been ingested.
Trough levels were measured approximately 48 hours later.
[0058] Compliance with the diet was monitored with a seven day
dietary recall questionnaire directed at calcium rich foods. Daily
calcium intake was estimated by adding the calcium content of
calcium rich foods identified by the questionnaire to the estimated
calcium content of the basal diet. The calcium content of the basal
diet was calculated to be 1 mg of calcium/9 Kcal. Caloric intake
was estimated with the use of the Food Processor 7.0 software (ESHA
Research, Salem, Oreg.).
[0059] After completing the four week treatment period, patients
were monitored for four additional weeks. Serum calcium was checked
in weeks 5 and 7 and tumor measurements were obtained in week 7.
All toxicities were graded according to NCI Common Toxicity
Criteria. Response was assessed according to WHO guidelines.
[0060] Statistical Considerations
[0061] The planned dose escalation was governed by the multistage
escalation scheme described by Gordon and Willson, 1992. The
maximal tolerated dose (MTD) was defined as that dose at which 1/3
or fewer of the patients experienced Grade 3 toxicity (64).
Patients who had evidence of response or stable disease, and no
Grade 3 or greater toxicity were permitted to reenroll and receive
either the same dose or the next higher dose of calcitriol.
Statistical analysis was performed using StatView 5.0 for Windows
software (SAS Institute, Cary, N.C.)
RESULTS
[0062] Fifteen different patients were enrolled in 20 cycles of
therapy (Table 2). Two patients were withdrawn from the study prior
to completion of the four week regimen because of disease
progression. No patient withdrew because of toxicity of therapy or
unacceptability of the diet. Five patients reenrolled for a second
cycle of treatment.
2TABLE 2 Individual Patients Enrolled on Study Cycle 1 dose Cycle 2
dose Patient Age Gender Malignancy (.mu.g/kg) (.mu.g/kg) 1 79 male
Adenocarcinoma of the prostate 0.06 0.12 2 42 female Adenocarcinoma
of the breast 0.12 3 70 male Adenocarcinoma of the lung 0.24 4 72
male Melanoma 0.48 5 53 male Squamous Cell of the tonsil 0.48 6 48
female Hepatocellular carcinoma 0.80 1.60 7 80 male Adenocarcinoma
of the prostate 0.96 2.00 8 53 female Adenocarcinoma of the breast
1.60 9 77 female Adenocarcinoma of the lung 1.92 2.00 10 78 male
Adenocarcinoma of the prostate 2.00 11 69 male Adenocarcinoma of
the prostate 2.00 12 46 female Adenocarcinoma of the breast 2.00 13
47 female Gastrointestinal stromal tumor 2.00 14 71 male
Adenocarcinoma of the pancreas 2.80 2.80 15 76 male Adenocarcinoma
of the prostate 2.80
[0063] No deaths occurred. No patient withdrew from the study due
to toxicity, and no Grade 3 or higher toxicity was seen. All
observed toxicities are listed in Table 3.
3TABLE 3 Toxicities developed during each treatment course (N = 20)
Toxicity Grade 1 Grade 2 Grade 3 Grade 4 Leukopenia .sup. 5.sup.1
.sup. 1.sup.2 0 0 Anemia 3 .sup. 4.sup.2 0 0 Thrombocytopenia 2 0 0
0 Hypercalcemia 8 0 0 0 Creatinine elevation 4 0 0 0 Bilirubin
elevation 2 0 0 0 ALT elevation 1 0 0 0 Alkaline phosphatase
elevation 2 .sup. 1.sup.2 0 0 Nausea and vomiting 5 2 0 0 Diarrhea
3 1 0 0 Constipation 5 0 0 0 Dyspepsia 4 0 0 0 Headache 5 0 0 0
Fever 2 0 0 0 Skin rash 1 0 0 0 Bone or muscle pain 8 0 0 0
.sup.1All were within normal limits of our laboratory (3.4-10.0
k/mm.sup.3) but fell into the Grade 1 toxicity range of 3.0-3.9
k/mm.sup.3 .sup.2All had Grade 1 abnormalities prior to entry onto
study.
[0064] The normal range for serum 1,25-dihydroxyvitamin D levels is
0.05-0.16 nM (20-65 pg/ml). An approximately linear increase in the
peak level was observed with increasing dose until the 0.48
.mu.g/kg dose (Table 4, FIG. 1). Above this dose, a further
elevation of peak levels was not seen. Serum 1,25-dihydroxyvitamin
D trough levels returned to normal or near normal levels by 48
hours (FIG. 1). A limited study of calcitriol pharmacokinetics
showed the expected decay in 1,25-dihydroxyvitamin D levels after
hour 6 (FIG. 2).
4TABLE 4 Mean Peak and 48 hour 1,25-dihydroxyvitamin D levels by
dose Dose level (.mu.g/kg) Patients Mean peak (nM) Mean 48 hour
level 0.06 1 0.71 0.27 0.12 2 1.10 0.14 0.24 1 2.27 0.21 0.48 2
4.11 0.23 0.80 1 3.53 0.96 1 3.83 1.60 2 3.65 1.92 1 3.34 2.00 6
4.07 0.26 2.80 2 2.96
[0065] Mean serum calcium (normal range 8.5-10.5 mg/dL) increased
from 9.55 (SD 0.57) mg/dL prior to treatment to 9.76 (SD 0.63)
mg/dL 24 hours later and to 9.88 (SD 0.68) mg/dL at 48 hours
(p=0.0002 by a two way repeated measures analysis of variance). All
calcium levels above the normal range returned to normal within 2
days with no intervention. Mean serum phosphate (normal range
2.2-4.2 mg/dL) increased from 3.43 (SD 0.56) mg/dL prior to
treatment to 3.98 (SD 0.57) mg/dL 24 hours later and dropped to
3.86 (SD 0.53) mg/dL at 48 hours (p<0.0001 by a two way repeated
measures analysis of variance). Mean 24 hour urinary calcium
excretion (normal range 100-300 mg) increased from 130 (SD 62) mg
with a range of 44-292 mg prior to treatment to 263 (SD 125) mg
with a range of 59-594 on treatment, measured on day 2 of each
treatment week (p<0.0001 by a one way repeated measures analysis
of variance). There was no statistically significant increase in
urinary calcium excretion during the treatment period by the
Bonferroni/Dunn test.
[0066] Five of eight patients with measurable disease had stable
disease. Among them, an adenocarcinoma of the lung patient, an
adenocarcinoma of the pancreas patient, and a hepatocellular
carcinoma patient received two cycles of therapy and remained
stable for the entire 16 weeks of their time on study. The
hepatocellular carcinoma patient had an associated 70% decline in
her serum AFP level. The remaining three patients with measurable
disease had evidence of progressive disease.
[0067] Four of seven patients without measurable disease had no
evidence of progression. Among them was the breast cancer patient
described in Example 3. A prostate cancer patient received two
cycles of therapy, and had a stable PSA for the entire 16 weeks
during which the drug was administered, in spite of a rapidly
rising PSA prior to enrollment. The remaining three patients
without measurable disease had either tumor marker or clinical
evidence of progressive disease.
[0068] No patient developed dose-limiting hypercalcemic toxicity
from calcitriol (<2 mcg/day). Measurements of peak blood
calcitriol levels in patients indicate that blood levels (up to 8.9
nM) are at a level known to be growth inhibitory for cancer cells
in culture. Furthermore, the drug calcitriol is essentially
completely cleared from the blood by day 3, and this rapid
clearance explains the increased safety profile of the weekly pulse
schedule.
[0069] An unanticipated result was the finding that escalation of
calcitriol dose beyond dose level 0.48 .mu.g/kg did not result in
further increases in peak calcitriol levels. More detailed
measurement of calcitriol levels in one patient (dose level 2.0
.mu.g/kg) indicated that absorption is saturated at high doses
rather than delayed, as neither the peak levels of calcitriol are
delayed and the half-life of the drug is not extended beyond the
usual time observed in lower dose studies. The maximal tolerated
dose (MTD) of calcitriol was not determined by the data presented
in this example.
[0070] In summary, pulsed weekly administration of calcitriol
allows substantial escalation of the total weekly dose of
calcitriol that can be administered to patients with advanced
malignancies. Peak blood levels of calcitriol about 25 fold above
the upper limit of normal are achieved with minimal toxicity. These
levels are well into the range where antiproliferative effects of
calcitriol are observed. Based on the observation that blood levels
of calcitriol do not increase linearly with increased dose beyond
the 0.48 .mu.g/kg level, a dose level of 0.5 .mu.g/kg is an example
of a dose that is therapeutically effective in patients whose tumor
responds to this therapy, but which does not result in unacceptable
hypercalcemia.
EXAMPLE 6
Preparation of Pharmaceutical Dosage Forms
[0071] Preparation of pharmaceutically acceptable compositions of
the Vitamin D drugs of the present invention can be accomplished
using methods well known to those with skill in the art. Any of the
common carriers such as sterile saline solution, plasma, etc., can
be utilized with the Vitamin D drugs of the invention. Routes of
administration include but are not limited to oral, intracranial
ventricular (icv), intrathecal (it), intravenous (iv), parenteral,
rectal, topical ophthalmic, subconjunctival, nasal, aural and
transdermal. The Vitamin D drugs of the invention may be
administered intravenously in any conventional medium for
intravenous injection such as an aqueous saline medium. Such medium
may also contain conventional pharmaceutical adjunct materials such
as, for example, pharmaceutically acceptable salts to adjust the
osmotic pressure, buffers, preservatives and the like. Among such
media are polysorbate, normal saline, lactated Ringer's solution,
and plasma. Vitamin D is somewhat insoluble, hence solubilizing
agents such as sesame oil, or equivalent lipophilic preparations,
may be used to administer the Vitamin D drug.
[0072] Embodiments of the invention comprising medicaments, such as
tablets or capsules, can be prepared with conventional
pharmaceutically acceptable carriers, adjuvants and counterions as
would be known to those of skill in the art. The medicaments are
preferably in the form of a unit dose in solid, semi-solid and
liquid dosage forms such as tablets, pills, powders, liquid
solutions or suspensions, and injectable and infusible solutions,
for example a unit dose vial. Effective dosage ranges included in
the unit dose for calcitriol vary from about 5 mcg to about 100
mcg. The unit dosages of the clacitriol are much higher than
previously used, because of the unanticipated finding that high
pulse, therapeutically effective doses of the drug can be given
without inducing symptomatic hypercalcemia.
EXAMPLE 7
Determining Binding Affinity
[0073] Binding affinity of the Vitamin D drugs for the Vitamin D
receptor can be determined by any acceptable means, such as the VDR
binding analysis and Scatchard plots in Peehl et al., Cancer
Research 54:805-810, 1994., which is incorporated by reference.
[0074] VDR affinity can be assayed by a competitive receptor assay
with radio-labeled calcitriol to determine the Relative Competitive
Index (RCI) wherein the RCI for calcitriol is set at 100. The RCI
of some of the Vitamin D analogs is set forth in Appendix 3.
EXAMPLE 8
Detecting Vitamin D Receptor on Tumor Cells
[0075] The presence of the VDR on tumor cells can be detected by
the methods set forth in Peehl et al., which has been incorporated
by reference in Example 7. A variety of other assays can be used to
detect the VDR, including immunohistochemistry (Kaiser et al., J.
Cancer Res. Clin. Onc 122:356-359, 1996); Western blot (Cross et
al., Anticancer Research 16:2333-2338, 1996); ligand binding assays
and DNA probe hybridization to RNA (Northern blot) (Endocrinology
132:1952-1960); and detection of RNA by ribonuclease protection
assay (Shabahang et al., Annals of Surg. Onc. 3:144-149, 1996).
[0076] In view of the many possible embodiments to which the
principles of our invention may be applied, it should be recognized
that the illustrated embodiment is only a preferred example of the
invention and should not be taken as a limitation on the scope of
the invention. Rather, the scope of the invention is defined by the
following claims. We therefore claim as our invention all that
comes within the scope and spirit of these claims.
[0077] The numerical values in the table for Relative Competitive
Index (RCI) for the nuclear VDR and the RCI for the vitamin D
binding protein (DBP) tabulate the data for the indicated parameter
in relation to the result for 1.alpha.,25(OH).sub.2D.sub.3 which is
normalized to 100; thus the data values represent percentages of
the 1.alpha.,25(OH).sub.2D.sub.3 value. The key to the data entries
is as follows. The RCI measures the relative ability of an analog
under in vitro conditions to compete with
[3H]1.alpha.,25(OH).sub.2D.sub.3 for binding to the nuclear
1.alpha.,25(OH).sub.2D.sub.3 receptor (VDR) (141) or the plasma
vitamin D transport protein (DBP) (142). The cell differentiation
data are related to that for 1.alpha.,25(OH).sub.2D.sub.3 for which
the value is set at 1.00. The calcemic index data are set at a
value of 1.00 relative to that of 1.alpha.,25(OH).sub.2D.sub.3.
[0078] SPC indicates the species of origin of the nVDR where
c=chick intestine, r=rat intestine, b=bovine thymus, p=pig
intestine, m=MCF-7 human breast cancer cells, z=rat osteosarcoma
ROS 17/2.8 cells. For the DBP, all data are obtained from the human
protein. The cell differentiation data are obtained in human
transformed cell lines where h=HL-60 cells, u=U-937. The calcemic
index is a measure of the relative ability of an analog to generate
a "calcemic" response, which is defined differently depending upon
the assay being conducted; c or i=a measure of intestinal Ca.sup.2+
absorption (ICA) or bone Ca.sup.2+ mobilizing activity (BCM) in the
vitamin D-deficient chick (146,553); b=bone resorption;
cbp=induction of the vitamin D-induced calcium binding protein,
calbindin-D.sub.28k; sc=elevation serum Ca.sup.2+=concentrations- ;
oc=an increase in serum osteocalcin levels; r=an increase in
urinary Ca.sup.2+ excretion. Reference numbers are indicated in
parentheses, and refer to regerence numbers in Boleillon et. al,
1995.
* * * * *