Preventing Metabolic Side Effects of Thiazide Diuretics With KMgCitrate
Phase: NA
Conditions: Hypertension
Sponsor: University of Texas Southwestern Medical Center

1 
 
“Protocol Amendment: New Protocol”: IND 116,208 KMgCit Powder 
IRB Study #: ---; August 31, 2015 [F]  
 
Study Protocol 
7.1. Title of Project: Amelioration of Chlorthalidone-Induced Hypokalemia and 
Metabolic Disturbances by Potassium-Magnesium Citrate (KMgCit) Powder 
Principal Investigator: Wanpen Vongpatanasin, M.D., Hypertension Section, 
Division of Cardiology, Department of Internal Medicine, University of Texas 
Southwestern Medical Center (UTSW) 
Co-Investigator: Orson W. Moe, M.D., Center for Mineral Metabolism and Clinical 
Research, UTSW 
 
In a just completed study (IND 116,208; IRB Study #: STU 072012-001), 
potassium magnesium citrate (KMgCit) powder did not alter blood pressure in 
patients with pre- and Stage I hypertension, while potassium chloride (KCl) 
powder lowered it. However, KMgCit powder reduced urinary 8-isoprostane 
serum FGF23. Chlorthalidone (CTD), a thiazide diuretic widely used for 
treatment of hypertension, produces certain metabolic disturbances associated 
with Type II diabetes and metabolic syndrome. In this “protocol amendment: new 
protocol”, these circumstances led us to explore whether KMgCit powder co-
administered with CTD would correct hypokalemia, and ameliorate metabolic 
disturbances of CTD therapy among patients with Stage I hypertension. We shall 
examine the effect of CTD alone, CTD with KMgCit powder or CTD with KCl 
powder in a parallel design.  
1. Specific Aims: 
  
Aim 1. To compare the effect of KMgCit powder and KCl powder on correction or 
prevention of CTD-induced hypokalemia.  
Aim 2. To examine CTD-induced changes in renin-angiotensin aldosterone (RAA) 
system, oxidative stress, and insulin sensitivity between KMgCit powder and KCl 
powder supplementation.  
Aim 3. To evaluate hepatic fat accumulation by MRS.  
Aim 4. To compare the Mg status from measurement of muscle Mg by MRS 
between CTD + KMgCit powder and CTD + KCl powder.  
 
It is expected that: (a) KMgCit powder would be equally effective in 
correcting/preventing CTD-induced hypokalemia as KCl powder, without 
incurring hypochloremic metabolic alkalosis. (b) Compared to KCl powder plus 
CTD, KMgCit powder added to CTD would reveal lower serum aldosterone, 
urinary isoprostanes, serum glucose, and HOMA-IR and serum FGF23. (c) 
Hepatic fat content would increase on CTD plus KCl powder, but would decline 
on CTD + KMgCit powder. (d) Muscle Mg would increase on KMgCit powder + 

2 
 
CTD, but would be decreased on KCl powder + CTD. It is hypothesized that CTD 
produces various metabolic disturbances that interact with each other or act 
independently to alter the propensity for Type II diabetes and metabolic 
syndrome, and that KMgCit powder ameliorates these disturbances or 
propensity. This protocol will put a special focus on testing a scheme that links 
Mg depletion from CTD to hepatic fat accumulation and insulin resistance.   
2. Background: 
  
The idea for this study came from a just completed trial (IND 116,208 
KMgCit Powder; IRB #STU 072012-001). We examined whether KMgCit powder 
could lower blood pressure in patients with pre- and Stage I hypertension, by 
providing three components of the “DASH” diet (potassium, magnesium and 
alkali) that have been shown to lower blood pressure (Sacks, 2001). KMgCit 
powder was compared with potassium citrate (KCit) powder, KCl powder and 
placebo. KMgCit powder was shown to be ineffective in lowering blood pressure.   
 
However, the above study disclosed that KMgCit powder might have other 
usefulness, since it seemed to ameliorate some of the metabolic disturbances 
associated with CTD.  
Metabolic disturbances of CTD 
 
CTD is a thiazide (TZ) type diuretic advocated by many guidelines for the 
treatment of hypertension (NICE 2011), owing to its longer half-life and greater 
antihypertensive efficacy at the clinically recommended doses than other 
thiazide diuretics (Vongpatanasin, 2015). Despite these favorable properties and 
increasing popularity, CTD and other TZ diuretics are known to cause various 
metabolic disturbances, such as hypokalemia, activation of renin-angiotensin 
system and sympathetic nervous system (RAA-SNS) (Menon, 2009), oxidative 
stress (Ribeiro, 2013; Reungjui, 2007), dyslipidemia (Eriksson, 2008; Price, 
2013), increased FGF23 synthesis (Pathare, 2012), insulin resistance (Xie, 2005; 
Price, 2013; Raheja, 2012; Menon, 2009), and Mg depletion (Sheehan, 1982; 
Hollifield, 1987).   
Mechanism for various metabolic disturbances of CTD/TZ ultimately 
contributing to the development of type II diabetes and metabolic syndrome are 
unknown. The traditional explanation implicating hypokalemia has been 
questioned.  
  
CTD/TZ-induced potassium depletion and hypokalemia are thought to be 
the main cause of hyperglycemia by impairing insulin secretion. However, serum 
insulin increases rather than decreases with TZ diuretics (Carter 2008, Raheja 
2012). The maintenance of normokalemia during CTD administration by K 
supplementation failed to prevent CTD-induced increase in fasting plasma 
glucose and HOMA-IR, an index of insulin resistance (Raheja, 2012; Menon, 
2009). CTD is known to induce a sustained activation of RAA-SNS in 

3 
 
hypertensive patients (Menon 2009, Raheja 2012). This neurohormonal 
activation is independent of serum potassium as it is unaffected by K 
supplementation (Menon 2009, Raheja 2012). Thiazide diuretic has been shown 
to increase renal oxidative stress, which cannot be accounted for by the presence 
of hypokalemia alone (Reungjui, 2007).  
  
Various metabolic disturbances of CTD/TZ might be pathogenetically 
linked, involving a pathway independent of hypokalemia. Some metabolic 
disturbance might directly cause insulin resistance and Type II diabetes. Other 
factors might do so indirectly by affecting other metabolic disturbances.  
Activation of RAA system has also been implicated in the pathogenesis of 
insulin resistance by inhibiting insulin signaling pathway in the adipocytes and 
skeletal muscle (Wada, 2009; Hitomi, 2007), resulting in impaired insulin 
mediated glucose uptake (Luther, 2011; Selvaraj, 2009). The deleterious effect of 
RAA system activation is mediated at least in part by increased oxidative stress 
(Sowers, 2009). Activation of the SNS system may further promote insulin 
resistance by reducing skeletal muscle glucose uptake both by flow-dependent 
(Jamerson, 1993; Sartori, 1999) and flow-independent (Lembo, 1994; 
Navegantes, 2003) mechanisms. 
  
Mg depletion from CTD/TZ may cause renal potassium loss and refractory 
hypokalemia (Whang, 1977). Magnesium supplementation alone without 
potassium ameliorated TZ-induced hypokalemia (Ruml, 1999). Activation of RAA 
system by CTD or TZ may contribute to Mg depletion. Inverse correlation between 
plasma renin activity and serum Mg has been demonstrated in a cross sectional 
study of patients with untreated essential hypertension (Resnick, 1983). In 
normal volunteers, serum aldosterone at baseline and after angiotensin II 
infusion was significantly higher on Mg-deficient diet than on Mg-replete diet 
(Nadler, 1993). Co-infusion of Mg with angiotensin II attenuated the rise in 
serum aldosterone, suggesting direct inhibitory effect of Mg on the RAA system. 
KMgCit powder as a countermeasure for metabolic disturbances of CTD 
Effective strategy in preventing the metabolic disturbances of CTD/TZ has 
not been fully developed.  
  
KCl supplementation is ineffective. Correction of hypokalemia by KCl did 
not reverse CTD-induced fasting hyperglycemia and increased HOMA-IR (Raheja, 
2012; Menon, 2009). CTD-induced activation of RAA-SNS was unaffected by KCl 
supplementation (Menon 2009, Raheja 2012).  In TZ-treated patients with 
hypertension, KCl supplementation did not reduce serum aldosterone (Kaplan, 
1985).  
  
KMgCit powder possesses distinct properties apart from KCl powder in 
overcoming deleterious metabolic disturbances of CTD therapy. Considerable 
data on physiological-clinical action of this drug have been obtained by this 

4 
 
investigative group using a tablet formulation (under IND 32,284 & 36,276; NDA 
20-964), as summarized below.     
 
Physiological properties of KMgCit. This preparation was shown to confer 
equivalent potassium bioavailability as potassium chloride, similar magnesium 
bioavailability as magnesium citrate, and greater alkali load than potassium 
citrate (Koenig, 1991). In patients with calcium nephrolithiasis, KMgCit 
significantly increased urinary pH and citrate compared to placebo. It was more 
effective than potassium citrate in inhibiting the crystallization of uric acid and 
calcium oxalate in urine (Pak, 1992). 
  
Some of the orally administered citrate may appear in urine by escaping 
hepatic metabolism in vivo. Serum citrate concentration increased significantly 
within 30 minutes after a single oral dose of citric acid and remained significantly 
elevated for 3 hours after citric acid load. Commensurate with this change, 
urinary citrate excretion peaked at 2 hours and gradually decreased during the 
next 2 hours after a citric acid load (Sakhaee, 1992).  
  
KMgCit was equally effective as potassium chloride in correcting HCTZ-
induced hypokalemia. In addition, KMgCit, but not KCl, produced a small but 
significant increase in serum magnesium concentration by delivering a 
magnesium load, and it conferred alkalinizing and citraturic actions (Pak, 2000).  
  
The above actions of KMgCit – potassium and magnesium load, citraturic 
action, and alkali load – might have a role in mitigating metabolic disturbances 
of CTD, as discussed below.   
  
Prevention of TZ-induced hypokalemia by KMgCit from provision of K and 
Mg load. In healthy subjects taking HCTZ, the addition of KMgCit 
corrected/prevented hypokalemia (Ruml, 1999; Wuermser, 2000), without 
producing metabolic alkalosis (Odvina, 2006).  
  
Potential inhibition of RAA-SNS activation by KMgCit powder from provision 
of Mg.  Pilot data from our laboratory indicated that magnesium supplementation 
attenuated sympathetic mediated vasoconstriction during muscle contraction in 
spontaneously hypertensive rats. Normally, exercise triggers parallel increases 
in sympathetic nervous system to the resting and exercising skeletal muscle. In 
the 
resting 
muscle, 
sympathetic 
activation 
triggers 
alpha-adrenergic 
vasoconstriction, thereby redirecting blood flow to the metabolically active 
muscles. In the working muscles, sympathetic vasoconstriction is markedly 
blunted by local vasodilator metabolites, particularly nitric oxide. This 
phenomenon, known as functional sympatholysis, is thought to be a protective 
mechanism to optimize blood flow to the working muscle (Remensnyder JP, 
1962). Our study indicated that functional sympatholysis is impaired in 
hypertensive 
patients, 
which 
may 
contribute 
to 
exercise 
intolerance 
(Vongpatanasin 2011). The impairment in functional sympatholysis in 

5 
 
hypertension was shown to be related to increased skeletal muscle oxidative 
stress that inactivates NO (Zhoa 2006).  
In 
our 
study 
in 
spontaneously 
hypertensive 
rats, 
dietary 
Mg 
supplementation blunted sympathetic mediated vasoconstriction (A, right) 
compared with Mg deficient diet (A, left), and reduced muscle oxidative stress 
both in contracting muscle and resting muscle (B) (Fig. 1). 
A. 
 
 
 
 
 
 
 
B. 
 
 
 
 
  
 
 
 
 
 
Fig. 1: A. percent change in vascular conductance (VC) in the femoral artery in response 
to lumbar sympathetic nerve stimulation (1, 2.5, and 5 Hz) at rest (white bars) and 
during muscle contraction (grey bars) in spontaneously hypertensive rats (SHRs) treated 
with Mg deficient diet (left, 30 mg/kg) vs. high Mg diet (4.8 grams/kg, right) for 12 weeks. 
Sympathetic mediated reduction in vascular conductance is attenuated in the SHRs 
treated with high Mg diet compared to Mg deficient diet, B. Summary data showing 
increased oxidative stress as evidenced by increased ethidium fluorescence (DHE/DAPI 
ratio) in resting and contracting muscles of SHRs treated with Mg deficient diet 
compared to high Mg diet.  
   
Potential inhibition of oxidative stress by KMgCit powder from provision of 
citric acid, alkali and magnesium. A high dose of citric acid (1–2 g/kg) reduced 
brain lipid peroxidation and inflammation, liver damage, and DNA fragmentation 
in mice exposed to lipopolysaccharide, suggesting a protective role of citric acid 
in endotoxin-induced oxidative stress (Abdel-Salam, 2014).  
Supporting an inhibitory role of alkali, a calcium/potassium salt of 
hydroxycitric acid (rich in alkali) extracted from the dried fruit rind of the plant 
Garcinia cambogia reduced inflammation, oxidative stress, and insulin 
resistance in obese Zucker rats (Asghar, 2007). In the Dietary Approaches to 
Stop Hypertension (DASH/SRD) trial, a diet rich in alkali and magnesium and 
low in sodium was shown to reduce urinary F2-isoprostanes by 31% in 
hypertensive patients with heart failure with preserved ejection fraction, 
suggesting decreased oxidative stress (Hummel, 2012). Consumption of black 
currant juice was shown to reduce plasma F2-isoprostanes and improve 
endothelial function in individuals with habitually low dietary intake of fruits 

6 
 
and vegetables (Khan, 2014). Induction of metabolic acidosis with oral 
ammonium chloride for 5 days in normal volunteers increased leucine 
appearance from body protein and led to leucine oxidation, suggesting increased 
protein degradation and amino acid oxidation (Reaich, 1992). It can be inferred 
that alkali load by KMgCit powder would have an opposite effect.  
As evidence of its protective role, Mg was shown to ameliorate creatine 
kinase-MB elevation, oxidative stress, lactate accumulation, and pyruvate 
reduction, as well as preserve creatinine phosphate, adenine nucleotides and 
Na(+),K(+)-ATPase 
activity, 
in 
mice 
with 
lipopolysaccharide-induced 
cardiotoxicity (Ahmed, 2012). Dietary magnesium restriction increased 
ischemic/reperfusion injury in the isolated rat hearts. Dietary Mg deficiency was 
shown to promote cardiomyopathy, in situ cardiac dysfunction, and myocardial 
intolerance to secondary stresses. 
Significant protection against most of 
these Mg deficiency-mediated events 
has been observed with interventions 
that modulate neuronal substance P 
release or its bioactivity, and with 
several 
antioxidants 
(vitamin 
E, 
probucol, epicaptopril, d-propranolol) 
(Kramer, 2009). 
 
We obtained the most direct 
evidence 
of 
KMgCit 
powder’s 
inhibition of oxidative stress in a just 
completed 
trial 
(original 
IND 
116,208). Thirty patients with pre- or 
Stage I hypertension underwent a 
crossover trial, whereby they took 
KMgCit powder, KCit powder, KCl 
powder or placebo for 4 weeks. The 
potassium content during the K salt 
phases was 20 meq bid. In the KMgCit powder phase, patients also took 10 meq 
Mg bid. A spot urine sample was obtained at the end of 4 weeks of treatment of 
each phase. Urinary 8-isoprostane was significantly lower during the KMgCit 
powder phase than during other phases (Fig. 2).  
  
Potential amelioration of dyslipidemia by KMgCit powder via Mg load. Mg 
supplementation with bittern, a natural MgCl2 solution from sea or salt lake 
water, reduced the appearance in serum of lipid components after a fat load in 
normal volunteers (Kishimoto, 2010). The concentrations of apo-B48, remnant-
like particle cholesterol, and nonesterified fatty acid were significantly lower at 2 
hours after the fat-with-Mg meal compared with the fat-only meal.  Thus, Mg 
supplementation by KMgCit powder could inhibit fat absorption and improve 
postprandial hyperlipidemia.   
Fig. 
2. 
Spot 
urinary 
8-isoprostane 
following 4 weeks of treatment with 
KMgCit powder, KCit powder, KCl powder 
or placebo. ** p < 0.05 and † < 0.001 from 
the KMgCit powder. p = 0.11 between KCit 
powder and placebo.   

7 
 
  
In rabbits fed a high cholesterol diet (1% or 2%), oral magnesium aspartate 
hydrochloride: (i) lowered serum cholesterol and triglyceride in normal (25-35%) 
as well as atherosclerotic (20-40%) animals, and (ii) attenuated the 
atherosclerotic process markedly. In addition, dietary deficiency of Mg 
augmented atherogenesis markedly and stimulated (or activated) macrophages 
in the reticuloendothelial system (Altura, 1990; Smith 2013). In rats, Mg 
deficient diet reduced post-heparin lipase activity and hepatic lipase, resulting 
in increased serum triglycerides and triglyceride content of chylomicrons, very 
low density lipoproteins, low density lipoproteins and high density lipoproteins 
(Rayssiguier, 1991).  
  
Potential suppression of FGF23 synthesis by KMgCit powder from alkali and 
Mg load.  In mouse calvarial and osteoblast preparations, a culture medium 
mimicking metabolic alkalosis inhibited FGF23 protein and mRNA (Krieger, 
2012). Mice exposed to Mg-deficient diet showed a marked increase in serum 
FGF23 (van Angelen, 2013).  
  
Potential amelioration of insulin resistance and type II diabetes by KMgCit 
powder from alkali and Mg load. In a randomized crossover study in normal 
volunteers, Mg deficient diet reduced insulin sensitivity (estimated from 
intravenous glucose tolerance test), compared to Mg repleted diet (Nadler 1993). 
In a double-blind, placebo-controlled study in prediabetic subjects, KCit 
(delivering alkali load) but not KCl reduced insulin resistance as revealed by 
HOMA-IR (Conen, 2015). In a cross-sectional study, Type 2 diabetes mellitus was 
associated with lower urinary pH and higher net acid excretion, independent of 
age, body weight, creatinine clearance, and dietary factors (Cameron 2006). 
Several populational studies have shown an association between dietary acid 
load and insulin resistance as well as the risk of type 2 diabetes mellitus (Akter 
2015 and Fagherazzi 2014). In the National Health and Nutrition Examination 
Surveys involving healthy US population, low serum bicarbonate and high anion 
gap were independently associated with increased plasma fasting insulin and 
triglyceride, suggesting increased risk of insulin resistance (Farwell 2008). These 
studies inferred potential protective action of KMgCit powder from alkali load.  
  
Several studies suggested a protective role of Mg. In a case control study 
of 163 women and 131 men, hypomagnesemia (serum magnesium ≤1.8 mg/dL) 
was independently associated with incident metabolic syndrome. Additional 
adjustment by serum C-reactive protein showed that metabolic syndrome 
remained associated with hypomagnesemia (OR 1.4; 95% CI 1.1-5.9) but not 
with oxidative stress determined by serum malondialdehyde concentration (OR 
1.1; 95% CI 0.9-5.9) (Guerrero-Romero, 2006). In a prospective observational 
study with median follow up of 16 years, high magnesium intake was a 
significant protective factor for the incidence of Type 2 diabetes, especially among 
subjects with insulin resistance, low-grade inflammation and a drinking habit 
(Hata, 2013).  

8 
 
  
In rats subjected to magnesium-deficient diet, hypomagnesemia had a 
deleterious effect on glucose metabolism by impairing both insulin secretion and 
action. Mg deficient rats displayed 45% reduction in the glucose-stimulated 
insulin secretion. Mg restriction produced defective tyrosine kinase activity of 
insulin receptors, leading to decreased phosphorylation of the beta-subunit of 
the insulin receptor (Suarez, 1995). 
  
Potential 
Mg 
repletion 
by 
KMgCit 
powder from provision of 
bioavailable 
Mg. 
By 
providing bioavailable Mg, 
KMgCit 
powder 
should 
prevent/ameliorate CTD-
induced hypomagnesemia 
or Mg depletion. In prior 
studies 
with 
a 
tablet 
formulation of KMgCit, 30 
healthy 
subjects 
developed 
hypomagnesemia (serum 
Mg < 1.8 mg/dL) on HCTZ 
50 mg/day alone for 2-3 
weeks (Pak, 2000). When 
KMgCit 
was 
added 
to 
HCTZ treatment at a dose 
of 42 meq K and 21 meq 
Mg/day, 
serum 
Mg 
increased to the normal 
range 
(Fig. 
3).   
 
 
 
 
 
 
Among 131 subjects who took HCTZ, the percentage of subjects with 
hypomagnesemia progressively declined when KMgCit was added (Fig. 4, closed 
circles). However, the percentage of subjects with hypomagnesemia increased 
when KCl or potassium citrate was added to HCTZ treatment (open circles).   
Fig. 3. Effect of KMgCit on serum Mg concentration 
among 30 subjects who developed hypomagnesemia 
alone 
on 
HCTZ. 
KMgCit 
was 
added 
to 
hydrochlorothiazide (TZ) therapy for three weeks. † = p 
< 0.001.  

9 
 
 
Fig. 4. The percentage of subjects 
with 
hypomagnesemia 
during 
addition of KMgCit or KCit/KCl to 
ongoing HCTZ therapy. * p < 0.05, 
** p < 0.01 and † p< 0.001. Open 
symbols indicate within group 
comparisons, 
and 
symbols 
enclosed in parenthesis indicate 
between group comparisons. M1, 
M2 and M3 indicate 1, 2 or 3 
weeks of combined treatment.  
Summary 
  In 
summary, 
CTD 
may 
produce various metabolic disturbances, including hypokalemia, activation of 
RAA-SNS, oxidative stress, dyslipidemia, FGF23 synthesis, insulin resistance, 
and magnesium depletion (Fig. 5). These factors act solely or interact with each 
other to contribute to the development of Type II diabetes and metabolic 
syndrome. KMgCit powder can potentially overcome all of these metabolic 
disturbances. On the other hand, the action of KCl powder might be limited to 
correction of hypokalemia. 
 
Fig. 5. Scheme for CTD-induced metabolic disturbances that could lead to Type II 
diabetes and metabolic syndrome. These disturbances might be blocked by KMgCit 
powder.   
A special focus of this protocol will be on the CTD-induced Mg depletion 
(Fig. 6). CTD is well known to cause Mg depletion, hepatic steatosis and Type II 
diabetes-metabolic syndrome. We speculate that Mg depletion is responsible for 
hepatic fat deposition, which then produces insulin resistance. Co-
administration of KMgCit powder would avert Mg depletion, block hepatic fat 
deposition by restoring normal Mg status and direct intestinal binding of fat, 
thereby ameliorating insulin resistance. To test this hypothesis, we shall 
quantitate muscle Mg status and hepatic fat content by MRS.  
CTD
Type II Diabetes &
Metabolic Syndrome
Metabolic Disturbances
• Hypokalemia
• RAA-SNS activation
• Oxidative stress
• Dyslipidemia
• FGF23 synthesis
• Insulin resistance
• Mg depletion
KMgCit
Powder
Pre
TZ
M1
M2
M3
Percentage of Patients
with Hypomagnesemia
0
5
10
15
20
25
30
KMgCit
KCit or KCl
†
†
†
(*)
(**)
(**)

10 
 
 
Fig. 6. Hypothesis for the central role of Mg depletion and hepatic fat deposition in the 
development of CTD-induced metabolic disturbances.  
 
3. Significance: 
 
  
Thiazide type diuretics are widely used as a first line treatment of 
hypertension in a very large number of subjects (James 2014, Vongpatanasin 
2014). While this class of drugs is effective in lowering blood pressure, its use is 
complicated by various metabolic disturbances, many of which are features of 
metabolic syndrome induced or worsened by CTD. Some of these metabolic 
complications already coexist with hypertension and their exacerbation by TZ 
diuretics may not be readily recognized. KMgCit powder potentially represents a 
novel modality to not only prevent CD/TZ-induced hypokalemia, but also inhibit 
or ameliorate metabolic disturbances. This simple replacement of inorganic 
solutes can potentially allow CTD/TZ to exert its full antihypertensive efficacy 
without the negative drawback of worsening metabolic syndrome counteracting 
the benefits of blood pressure control.  
4. Preliminary Data/Past Work: 
  
This investigative group has already obtained preliminary data or 
published relevant work on all four aims of this proposal. Using a tablet 
formulation of KMgCit, the group has delineated physiological action of this 
compound in detail and some of its clinical effects. Safety of usage was also 
shown. Some data to be shown below were derived from original IND with KMgCit 
powder or in experimental animals.  
  
Effect of CTD on inflammation. In our pilot data in 16 subjects with 
uncomplicated stage I hypertension, serum highly sensitive C-reactive protein 
(hsCRP) rose significantly from 3.2 (1.0 – 4.6; 
median, 25th – 75th percentile) at baseline to 4.8 
(2.6 – 9.8) during CTD (p < 0.05) (Fig. 7).  In 
contrast, spironolactone (Spiro) had no effect on 
hsCRP levels in the same subjects.  
Fig. 7. Summary data showing changes in serum 
hsCRP (mcg/ml) in 16 untreated hypertensive 
subjects at baseline, after chlorthalidone treatment 
for 12 weeks, and after spironolactone treatment for 
12 weeks. 
CTD
Mg Depletion
Hepatic Fat
Deposition
Type II Diabetes &
Metabolic Syndrome
0
2
4
6
8
Baseline
CTD
Spiro
p < 0.05 vs baseline

11 
 
Correction/prevention of TZ-induced hypokalemia (Wuermser, 2000). After 
treatment with HCTZ 50 mg/day alone for 2-3 weeks, 30 patients took KMgCit 
tab 42 meq K and 21 meq 
Mg daily, while 30 subjects 
received KCl 42 meq/day, 
along with HCTZ for 3 
weeks 
(Fig. 
8). 
Serum 
potassium 
declined 
on 
HCTZ alone. It rose to the 
normal 
range 
on 
both 
KMgCit and KCl to an 
equivalent degree.  
 
Fig. 8. Effect of KMgCit and 
KCl on serum potassium. † 
represents p < 0.001 from the 
last week of HCTZ alone. 
 
  Inhibition of RAA-SNS 
activation. In spontaneous hypertensive rats, high Mg diet blunted sympathetic 
mediated vasoconstriction (see Fig. 1).  
Effect of KMgCit powder on oxidative stress. Urinary 8-isoprostane 
significantly declined on KMgCit powder. See Fig. 2.  
 
Effect of KMgCit powder on serum triglyceride. In a retrospective analysis 
of data from a prior study (Odvina, 2006), 8 patients took HCTZ 50 mg/day with 
KMgCit tab 40 meq K and 20 meq Mg/day for 6 months. 10 patients took HCTZ 
50 mg/day with KCl tab 40 meq K/day. Serum triglyceride declined by -32  41 
mg/dL from 1 month to 5 months of treatment in the KMgCit group, and 0  31 
in the KCl group (p = 0.09). Unfortunately, no baseline or 6 month data were 
available. 
 
Effect of KMgCit powder on serum FGF23. In 30 patients with pre- and 
Stage I hypertension participating in a crossover randomized trial, serum FGF23 
was significantly lower when they were taking KMgCit powder than KCl powder 
(46.1 +/- 16.7 Rel U/ml vs. 53.1 +/- 22.2 Rel U/ml, p=0.05).  
  
Prevention or correction of thiazide-induced hypomagnesemia. See Fig. 3 & 
Fig. 4.  
5. Methods of Procedure: 
Patients to be studied 
Serum potassium, mEq/L
2
3
4
5
6
KMgCit
KCl
†
†
†
†
†
†
†
†
Pre-treatment Last week
HCTZ
Week 1
supplement
Week 2
supplement
Week 3
supplement

12 
 
 
Studies will be conducted only in patients with treated or untreated stage 
I hypertension without concomitant cardiovascular diseases, chronic kidney 
diseases, or compelling indication for treatment with ACE inhibitor, beta 
blockers, or ARB. Patients will be adult men or women (> 21 years of age) of any 
ethnicity. Study is planned in 72 patients to account for potential dropout rate 
of 20%. 
Excluded will be patients with  
 diabetes mellitus, 
 Renal impairment (serum creatinine > 1.4 mg/dL), 
 Any heart diseases such as congestive heart failure, sustained arrhythmia, 
or coronary heart disease,  
 Chronic regular NSAID use,  
 Allergy to thiazide diuretics,  
 Gastro-esophageal reflux disease (GERD) requiring treatment with acid 
reducing agents or antacid more than once a week,  
 Esophageal-gastric ulcer or history of gastrointestinal bleeding,  
 Chronic diarrhea, vomiting, 
 Excessive sweating,   
 Unprovoked hypokalemia (serum K < 3.5 mmol/L) or hyperkalemia (serum 
K > 5.3 mmol/L),  
 Abnormal liver function test (AST or ALT above upper limit of normal 
range),  
 Subjects on any potassium supplement on a regular basis for any reason, 
such as patients with primary aldosteronism,  
 Pregnancy,   
 History of major depression, bipolar disorder, or schizophrenia,  
 History of substance abuse, 
 Gout,  
 Metabolic alkalosis, with serum bicarbonate > 32 meq/L,  
 Severe dietary salt restriction, less than1/2 spoonful or 50 meq 
sodium/day.  
 
Study design 
 
The patients will participate in a three-phase study in a parallel design 
(Fig. 9).  
 
 
 
 
  Fig. 9. Study scheme.  
 
 

13 
 
After baseline evaluation (Phase 1, without CTD), patients will take CTD 
alone for 2-3 weeks (Phase 2, CTD alone). They will then be randomized to two 
equal groups to take KMgCit powder or KCl powder along with CTD for 4 months 
(Phase 3).  
Test drugs 
  
KCl powder, made by and purchased from Sterling Pharmaceutical 
Services LLC, will contain 20 meq K as potassium chloride per sachet. KMgCit 
powder, also made by Sterling, will contain 20 meq K, 10 meq Mg, and 30 meq 
citrate, per sachet. These preparations will be prepared by the same method, and 
their composition will be the same, as those used in the completed trial (original 
IND 116,208).   
The sachets of KCl and KMgCit powder will be identical in appearance. 
During Phase 3, patients will dissolve the contents of each sachet of KCl or 
KMgCit powder in 250 ml water and ingest it with breakfast and again with 
dinner, to deliver 40 meq K per day. In the group taking KMgCit powder, patients 
will also receive Mg 20 meq and citrate 60 meq per day.   
  
A taste enhancer and sucralose will be added to improve tolerance of the 
test medications. Each sachet will be marked by a code, the identity of which 
will be known only to an independent study monitor. Each sachet will be labeled: 
Study drug # --; CTD-Induced Hypokalemia; Investigator: W. Vongpatanasin, 
M.D.  
 
Chlorthalidone, 25 mg generic tablets, will be purchased from a local 
pharmacy.   
Conflict of interest statement 
  
UT Southwestern had licensed KMgCit powder to Althos Pharma. 
Unfortunately, Althos Pharma is no longer operational due to financial 
difficulties. Accordingly, all the licensing agreements between UTSW and Althos 
Pharma, including one for KMgCit powder, have been terminated. (Tracy 
Roberson, licensing officer of UTSW’s Office of Technology Development, can be 
consulted for verification.) No relationship now exists between Althos Pharma 
and UTSW or investigators. 
 
Despite lack of industrial sponsorship, the investigators are convinced that 
the high scientific merit of the proposal mandates its pursuit. Thus, this protocol 
is investigator-initiated and research-driven, and not industry-sponsored. The 
initial costs of the study will be borne by the biotechnology program of CMMCR. 
CMMCR will purchase test medications from Sterling Pharmaceutical Services.  
 
Outline of the study (Table 1) 

14 
 
 
Baseline (Phase 1, time zero). At enrollment, various tests/procedures as 
outlined in Table 1 will be obtained. Patients who are on antihypertensive 
medications will be asked to stop them. 
CTD Alone (Phase 2). Patients will take CTD 25 mg/day. Two weeks later, 
they will return with a 24-h urine collection. Venipuncture and other procedures 
as outlined in Table 1 will be performed. Subjects who develop hypokalemia 
(serum K < 3.5 mEq/L) will immediately enter Phase 2. The remaining patients 
will continue on CTD for another week. On return at 3 weeks, all the 
tests/procedures will be performed in all patients.  
Table 1. Study Outline 
Weeks /months 
 
0 
2/3w   Rx 
1m 
2m 
3m 
4m 
 
Treatment 
 
 
 
   CTD  
 
 
   --------------------------------------------------- 
   KMgCit/KCl 
 
 
 
 ----------------------------------- 
Office blood pressure 
 
 
 
 
 
 
 
 
Serum tests  
 
 
 
 
 
 
 
 
 
 
Urine tests 
 
 
 
 
 
 
 
 
 
 
 
Side effects QN 
 
 
 
 
 
 
 
 
 
Diet history & BWt  
 
 
 
 
 
 
 
 
Sachet count  
 
 
 
 
 
 
 
 
 
CTD pill count 
 
 
 
 
 
 
 
 
 
Fecal occult blood  
 
 
 
 
 
 
 
 
 
Muscle Mg MRS 
 
 
 
 
 
 
 
 
 
Hepatic fat MRS 
 
 
 
 
 
 
 
 
_________________________________________________________________________________ 
Some patients may require only 2 weeks of CTD, instead of 3 weeks. Thus, the complete 
study duration might be shorter by 1 week. 
 
If patients develop symptomatic hypotension (blood pressure ≤ 90/60 
mmHg) after CTD treatment for 2 weeks, the dose of CTD will be reduced to 12.5 
mg daily temporarily. Blood pressure will be repeated 1 week after dose 
reduction. If BP is increased above 120/80 mmHg, the dose of CTD will be 
increased to 25 mg daily. Few patients who cannot later tolerate a higher dose 
of 25 mg/day will be withdrawn from the trial. If patients develop (BP ≥ 
130/80mmHg) after CTD treatment for 2 weeks, amlodipine 5 mg daily will be 
added to the regimen for blood pressure control. Blood pressure will be repeated 
in 1 week.  
 
 
 
CTD + KMgCit/KCl (Phase 3). Patients who developed hypokalemia at 2 
weeks of CTD will enter Phase 3. Those who completed three weeks of treatment 
with CTD alone will enter Phase 3 whether or not they develop hypokalemia. All 
patients will continue to take CTD 25 mg/day. All subjects will be randomized 
to receive KMgCit powder or KCl powder at a dose of 40 meq of K daily for 4 
months. Various tests will be performed at times indicated in Table 1.  

15 
 
 
Tests 
  
Office blood pressure. During clinic visits at baseline, at 2 and/or 3 weeks 
of CTD alone, at monthly intervals during combined treatment with CTD and 
KMgCit or KCl powder, the research nurse will obtain blood pressure by using 
the clinic’s oscillometric device (Welch Allyn, Vital Signs, WA). At each visit, three 
blood pressure measurements will be made after subjects sit quietly for at least 
10 minutes. The first reading will be discarded. The 2nd and 3rd measurements 
will be averaged.  
Serum tests. At baseline, 2 and/or 3 weeks after CTD alone, and at 2 and  
4 months of combined CTD and K salt treatment, a fasting venous blood sample 
will be obtained for electrolytes (potassium, sodium, chloride and carbon 
dioxide), magnesium, creatinine, calcium, phosphorus, albumin, triglyceride, 
cholesterol, LDL, HDL, and glucose (by Quest Laboratory). Serum PTH, insulin, 
8-isoprotanes, hsCRP, FGF23 and Klotho will be measured (by the Mineral 
Metabolism Laboratory, Moe’s Laboratory or O’Brien Center). At same times, a 
sample will be frozen for aldosterone (to be determined later when warranted by 
the results). Lymphocytes will be harvested from EDTA-treated blood sample and 
stored in RNALater for WNK1 mRNA (to be determined later when warranted in 
the laboratory of C.-L. Huang).   
 
Urine Tests. A 24-h urine sample will be collected under oil at baseline, 2 
and/or 3 weeks of CTD alone, 2 months and 4 months of combined treatment. 
Samples will be analyzed for stone risk factors (including potassium, pH, citrate, 
magnesium, sodium, calcium, phosphorus, sulfate, chloride, creatinine, and 
total volume), and net acid excretion by the Mineral Metabolism Laboratory. 
Samples will also be analyzed for 8-isoprostanes (by Moe’s laboratory) and frozen 
for aldosterone (to be determined later).   
 
At same times, a fresh spot urine sample will be obtained for creatinine 
(by Mineral Metabolism Laboratory), and frozen for WNK1 protein and mRNA, 
and exosomes (to be determined in the laboratories of Huang and Moe if 
warranted). WNK1 might be a mediator of changes in renal (sodium handling) 
and vascular (resistance) determinants of blood pressure that might ensue from 
varying potassium intake. Exosomes are shed membranes enclosing cytoplasm 
from the kidney shown to contain mRNA The measurement will allow us to 
assess levels of proteins and signaling molecules involved in the renal mineral 
transport.  
 
 Side effect questionnaire. At baseline, 2 and/or 3 weeks after CTD, and at 
monthly visits of combined treatments, the research nurse will complete the side 
effect questionnaire by interviewing the patients (Table 2). This history permits 
computation of “gastrointestinal symptom score”, a quantitative estimate of 
frequency and severity of gastrointestinal side effects (Ruml, 1999). 

16 
 
Other tests. At each follow-up visit, the research nurse (D. Arbique) will 
collect the test medications and count the number of unused packets or pills, in 
order to estimate compliance of patients in taking medications. At each visit, the 
research nurse will obtain history of unusual dietary intakes. Body weight will 
be measured. Smear of stool brought in by patients will be tested for occult blood.  
 
Muscle Mg by MRS. Intracellular muscle Mg will be obtained by magnetic 
resonance spectroscopy in the Department of Radiology.  
 
 
Hepatic fat by MRS. Hepatic TG content will be evaluated using a1.5 Tesla 
Gyroscan Achieva whole body clinical system (Philips Medical Systems, 
Cleveland, USA) equipped with software for localized spectroscopy as described 
(Price, 2013). In short, high-resolution morphological images are collected to 
serve as a “roadmap” for selection of a testing volume of 27 cc within the upper 
right hepatic lobe. All spectra are collected using PRESS sequence (Point 
RESolved Spectroscopy) for spatial localization and the signal acquisition with 
the following data acquisition parameters: Te = 27 ms, Tr = 3 s. All data are 
collected without water suppression. Areas of resonances from protons in water 
molecules and in methylenes of fatty acid chains will be evaluated with line-fit 
procedure using a commercial software (NUTS-ACORNNMR, Freemont, CA). 
 
Subjects with claustrophobia or metal devices precluding MRI, will not have MRI 
completed as part of this study. 
 
Table 2. Side Effect Questionnaire 
 
 
 

17 
 
6. Statistical Analysis 
Expectations (Table 3) 
 
 
 
 
Table 3. Anticipated Findings 
 
CTD CTD + KMgCit CTD + KCl 
Ks 
↓ 
= 
= 
Isoprostanes 
↑ 
↓ 
↑ 
Serum FGF23 = 
↓ 
= 
HOMA-IR 
↑ 
↓ 
↑ 
Aldosterone 
 
 
 
Muscle Mg 
 
 
 
Hepatic fat 
 
 
 
 
- 
On CTD alone, serum K would decline, reaching below normal in most 
patients. On both KMgCit powder and KCl powder along with CTD, serum 
K would increase to the normal limits. Similar levels of serum K would be 
observed during KMgCit powder vs. KCl powder.  
- 
Office blood pressure would decline during CTD alone. It would remain low 
on combined KMgCit powder or KCl powder with CTD.   
- 
It is expected that serum and urinary isoprostanes (marker of oxidative 
stress) would increase on CTD alone, decrease on combined KMgCit 
powder + CTD and remain unchanged on KCl powder + CTD.  
- 
We anticipate that serum FGF23 would increase on CTD alone, decrease 
on addition of KMgCit powder but not KCl powder.  
- 
Fasting plasma glucose and insulin would be higher during CTD when 
compared to baseline. KMgCit powder would attenuate the increase in 
plasma glucose and insulin, whereas KCl powder would not. HOMA-IR 
would show changes in the same direction.  
- 
Serum aldosterone (marker of renin-angiotensin system) would increase 
on CTD; the addition of KMgCit powder would attenuate this rise, but KCl 
powder would not.   
- 
The gastrointestinal symptom score during CTD alone will be similar to 
baseline. On combined treatment, gastrointestinal symptom score would 
be higher for KCl powder than KCit powder, with a larger percentage of 
patients complaining of dysgeusia on KCl powder.  
- 
At 3 weeks, 24-hour urinary potassium on KMgCit powder and KCl powder 
would be higher than CTD alone by about 35-40 meq/day. Urinary pH and 
citrate would be higher on KMgCit powder than CTD alone or KCl powder.  
- 
Free muscle Mg would increase on KMgCit powder but decline on KCl 
powder.  
- 
Hepatic fat would increase on KCl powder + CTD, but decline or remain 
unchanged on KMgCit + CTD.  
 
Sample Size Calculations 

18 
 
  
From our previous CTD studies, the mean increase in fasting serum  
glucose at CTD dose of 25 mg/day plus KCl supplementation to keep serum K 
constant was 10 mg/dL. The within subject SD of serum glucose was between 
2.0-8.5. Assuming SD = 8.5, power of 0.8 and  of 0.05, 60 patients (30 
randomized to KCl powder and 30 to KMgCit powder) would be needed to detect 
a potential reduction in serum glucose of 6 mg/dL by KMgCit. Assuming SD of 
5, power of 0.80 and  of 0.05, 60 patients would be needed to detect a potential 
reduction in serum glucose of 4 mg/dL by KMgCit. To account for a dropout of 
20%, we plan to recruit 72 subjects for the study. 
 
Statistical Methods 
  
Descriptive statistics and 95% confidence intervals will be used to 
summarize blood pressure and other responses for the three phases. Data from 
the three phases will be compared by using the repeated measures analysis of 
variance models. Significant differences between the three phases will be further 
analyzed by comparison of pairs of phases using contrasts constructed from the 
ANOVA models. By employing Bonferroni inequality to adjust for multiple 
testing, various comparisons between phases will be made. In addition, the 
existence of a trend in the differences of various parameters between phases will 
be sought (Perlman, 2002). Interim analysis will be performed at 50% completion 
of the trial. 
 
 

19 
 
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