25S,26-dihydroxycholecalciferol in the treatment of hypercalcitriolemic disease states

The invention is directed to a method in the treatment of hypercalcitriolemic disease states which comprises administering an effective amount of 25S,26-dihydroxycholecalciferol to a host in need of such treatment.

BRIEF SUMMARY OF THE INVENTION 
The invention is directed to a method for the treatment of 
hypercalcitriolemic disease states which comprises administering to a host 
in need of such treatment a therapeutically effective amount of 
25S,26-dihydroxycholecalciferol. 
DETAILED DESCRIPTION OF THE INVENTION 
The invention comprises a method for the treatment of hypercalcitriolemic 
disease states which comprises the administration of an effective amount 
of 25S,26-dihydroxycholecalciferol to a host in need of such treatment. 
25S,26-dihydroxycholecalciferol and its preparation are disclosed in 
Koizumi et al., J. Chem. Soc. Perkin Trans. I (1983) pages 1401-1410. 
25S,26-dihydroxycholecalciferol lowers endogenous levels of 
1.alpha.,25-dihydroxycholecalciferol, that is, it is hypocalcitriolemic. 
Accordingly, 25S,26-dihydroxycholecalciferol can be administered to warm 
blooded animals in dosages that are in the range of about 20 to about 1000 
micrograms/day for the treatment of hypercalcitriolemic disease states, 
such as, inoperative primary hyperparathyroidism, hypercalcemic 
granulomatous disease and certain forms of renal stone disease. 
25S,26-dihydroxycholecalciferol can be administered orally, subcutaneously, 
intramuscularly, intravenously or intraperitoneally. 
25S,26-dihydroxycholecalciferol can be formulated in compositions, such as, 
tablets, capsules, and the like or elixirs for oral administration or in 
sterile solutions or suspensions for parenteral administration. About 
20-1000 micrograms of 25S,26-dihydroxycholecalciferol can be compounded 
with a pharmaceutically acceptable vehicle, carrier, excipient, binder, 
preservative, stabilizer, flavor and the like in the unit dosage as called 
for by accepted pharmaceutical practice. The amount of active substance in 
the foregoing compositions or preparations is in the range previously 
indicated. 
Illustrative of the adjuvants which may be incorporated into capsules and 
the like are the following: a binder such as gum tragacanth, acacia, corn 
starch or gelatin; an excipient such as dihydroxyphosphate; a 
disintegrating agent such as corn starch, potato starch, algenic acid and 
the like; a lubricant such as magnesium stearate, a sweetening agent such 
as sucrose, lactose or saccharin; a flavoring agent such as peppermint, 
oil of wintergreen or cherry. Various other materials may be present as 
coating or to otherwise modify the physical form of a dosage unit. For 
instance, tablets may be coated with shellac, sugar or both. A syrup or 
elixir may contain the active compound, sucrose as a sweetening agent, 
methyl and propyl parabens as preservatives, a dye and a flavoring such as 
cherry or orange flavor. 
Sterile compositions for injection can be formulated according to 
conventional pharmaceutical practice by dissolving or suspending the 
active substance in a vehicle, such as water for injection, a naturally 
occurring vegetable oil, such as, sesame oil, coconut oil, peanut oil, 
cottonseed oil, and the like, or synthetic fatty vehicles such as, ethyl 
oleate or the like. Buffers, preservatives, antioxidants and the like can 
be incorporated as required. 
The useful activity of 25S,26-dihydroxycholecalciferol as an agent for the 
treatment of hypercalcitriolemic disease states can be demonstrated by the 
following test procedures. 
MATERIALS AND METHODS 
Animals 
Adult female Sprague-Dawley rats weighing between 190-200 g were used in 
all studies and were purchased from Harlan (Houston, TX). The animals were 
housed in the animal resource facility and were fed a standard vitamin 
D-replete rat chow diet (2.0% calcium and 0.9% phorphorus, Tekland, 
Winfield, OH) ad lib. After four to five days of equlibration the animals 
were divided into appropriate study groups and injected subcutaneously 
each day with various doses of 25(S),26(OH).sub.2 D.sub.3 or 
1,25(OH).sub.2 D.sub.3 metabolites. In some experiments, animals received 
both metabolites as two separate injections. Control animals received only 
vehicle (propylene glycol). 
Vitamin D Metabolite Assay 
Two ml of serum was obtained for each rat for measurement of 
25(S),26(OH).sub.2 D.sub.3, and 1,25-(OH).sub.2 D.sub.3. After spiking the 
samples with approximately 1200 cpm of radiolabeled 250HD.sub.3, 
24,25-(OH).sub.2 D.sub.3 and 1,25(OH).sub.2 D.sub.3 and allowing the 
samples to equilibrate for 30 minutes, isolation of the vitamin D 
metabolites was accomplished as described in Bishop JE, Norman AW, Coburn 
JW, Roberts PA, Henry, HL 1980 "Determination of the concentration of 
25-hydroxyvitamin D, 24,25-dihydroxyvitamin D and 1,25-dihydroxyvitamin D 
in a single two-milliliter plasma sample." Min Elect Metab 3: 181. Further 
separation and partial purification of the metabolites was effected with 
lipohilic Sephadex (Sigma, St. Louis, MO) liquid gel chromatography using 
a 9:1:1:0.015 (hexane:chloroform:methanol:H.sub.2 O) solvent system and a 
10.times.1 cm gel bed as previously described by Eisman JA, Hamstra AJ, 
Kream BD, DeLuca HF, 1975, "A sensitive, precise and convenient method for 
determination of 1,25-dihydroxyvitamin D in human plasma." Arch Biochem 
Biophys 176: 235. This chromatographic system effectively separates 250HD 
(5-11 ml) from the dihydroxy metabolites (14-30 ml). The trihydroxy 
metabolite 1,25,26(OH).sub.3 D is eluted from the column by changing the 
solvent system to 8:2:2:0.015 (hexane:chloroform:methanol:H.sub.2 O) after 
elution of the dihydroxy metabolites and collecting the next 12 ml. 
Final resolution and purification of the vitamin D metabolites was 
performed with high pressure liquid chromatography as previously described 
in Zerwekh et al., 1983, "Extra-renal production of 24,25-dihydroxyvitamin 
D in chronic renal failure during 25-hydroxyvitamin D.sub.3 therapy." Kid 
Int. 23: 401 and Pak et al., 1982, "Effects of short-term glucocorticoid 
administration in primary hyperparathyroidism: comparison to sarcoidiosis 
J. Clin. Endrocrinol Metab. 54: 824. 250HD was purified on a 5% 2-propanol 
in hexane solvent system. The three dihydroxyvitamin D metabolites were 
isolated and purified using an 11% 2-propanol in hexane solvent system. 
The assay of 25S,26-(OH).sub.2 D was performed using the rat serum vitamin 
D binding protein assay system described in Haddad JG, Chyu KF 1971 
"competitive protein binding assay for 25-hydroxycholecalciferol." J Clin 
Endocrinol Metab 33: 992. The assay of 1,25(OH).sub.2 D was accomplished 
using the chick intestinal receptor protein. 
TABLE 1 
______________________________________ 
Effect of 25S,26(OH).sub.2 D.sub.3 Administration 
on 1,25(OH).sub.2 D.sub.3 Mediated Increases in Serum 1,25(OH).sub.2 
D.sub.3 
25S,26(OH).sub.2 D 
1,25(OH).sub.2 D 
Exp. No. (ng/ml) (pg/ml) 
______________________________________ 
1. CONTROL 1.9 .+-. 0.2 
25 .+-. 3 
1,25(OH).sub.2 D.sub.3.sup.a 
3.0 .+-. 0.8 
206 .+-. 17 
(p &lt; 0.001) 
25S,26(OH).sub.2 D.sub.3.sup.b 
295 .+-. 7 59 .+-. 3 
+ (p &lt; 0.001) 
1,25(OH).sub.2 D.sub.3 
2. CONTROL 2.0 .+-. 0.2 
23 .+-. 3 
1,25(OH).sub.2 D.sub.3 
3.20 .+-. 0.3 
175 .+-. 52 
(p = 0.006) 
25S,26(OH).sub.2 D.sub.3 
168 .+-. 14 46 .+-. 4 
+ (p &lt; 0.01) 
1,25(OH).sub.2 D.sub.3 
3. CONTROL 1.20 .+-. 0.1 
27 .+-. 4 
1,25(OH).sub.2 D.sub.3 
0.9 .+-. 0.3 
180 .+-. 45 
(p &lt; 0.001) 
25S,26(OH).sub.2 D.sub.3 
236 .+-. 28 38 .+-. 6 
+ (p &lt; 0.006) 
1,25(OH).sub.2 D.sub.3 
______________________________________ 
Values in parenthesis represent p value of significant difference between 
dosed and control animals as determined by nonparametric analysis and 
Welch's approximation to the t test. All values expressed as mean .+-. SE 
for seven rats per group. 
.sup.a 1,25(OH).sub.2 D.sub.3 administered subcutaneously at a dose of 
0.15 .mu.g/d/rat for three days 
.sup.b 25S,26(OH).sub.2 D.sub.3 administered subcutaneously for seven day 
at a dose of 20 .mu.g/d/rat; on days 5, 6, and 7 both 25S,26(OH).sub.2 
D.sub.3 and 1,25(OH).sub.2 D.sub.3 (0.15 .mu.g/d/rat) were given as two 
separate injections. 
As the above results demonstrate, 25S,26-dihydroxycholecalciferol is active 
in reducing serum concentrations of 1,25-dihydroxycholecalciferol.