Introduction
Fragility fractures are common, with approximately 4.3 million new fractures in the European Union (EU) annually [
1]. Fragility fractures are associated with severe morbidity, increased mortality, high socioeconomic costs, and an increased risk of new imminent fractures [
1,
2]. The risk of imminent fractures could be effectively reduced with readily available preventative treatments [
3‐
5].
Denosumab is an antibody that effectively inhibits osteoclasts, resulting in increased bone mineral density and a reduced risk of fragility fractures [
4,
6]. Hypocalcemia is a known adverse event [
7‐
10], especially in patients with calcium-associated diseases, such as in secondary hyperparathyroidism, where the body is dependent on bone as a source of calcium to maintain adequate blood levels [
11].
The prevalence of hypocalcemia after denosumab injection was low in the original clinical trial (Fracture REduction Evaluation of Denosumab in Osteoporosis Every 6 Months, FREEDOM trial), at < 1% [
4,
6]; however, reports on real-world data indicate a higher prevalence of 4–26% [
7‐
10]. Methods to assess post-injection hypocalcemia vary, and in previous large-scale studies, calcium was not routinely analyzed directly after injections. Instead, study protocols included calcium analysis every 6 months, i.e., before injection and at predefined intervals (3, 6, 12, 24, and 36 months), or included any non-routine calcium value taken at predefined intervals [
4,
6‐
8,
10,
12]. These methods can miss the early transient post-injection hypocalcemia phase [
9,
13,
14]. Furthermore, non-routine calcium values might be biased in how the patients were selected for calcium analysis. These study designs could include a risk of both under- and overestimation of the actual prevalence.
In the clinical context, from 2011 until the introduction of the new guidelines in 2021, our local guidelines recommended to analyze ionized calcium (free bioactive calcium) before and 2 weeks after each denosumab injection for all patients, independent of known risk factors. In the present study, we analyzed the long-term real-world data of this unselected osteoporosis cohort.
The purpose of the study was to assess: 1) the prevalence of hypocalcemia after denosumab in a cohort routinely followed up with calcium measurements, 2) the risk factors for developing hypocalcemia, and 3) the risk factors for developing severe hypocalcemia.
Discussion
In the present study, using long-term real-world data on denosumab-treated osteoporosis patients followed up routinely regarding peri-injection calcium, we show that one in five patients had experienced post-injection hypocalcemia at least once during the treatment period. Most commonly, however, hypocalcemia was mild and not recurrent. Severe post-injection hypocalcemia, defined as an ionized calcium < 1.05 mmol/L, was seen after 1% of injections, of which a few patients needed inpatient care or an acute open care visit (n = 5). The risk factors for post-injection hypocalcemia were renal failure (CKD stage 4/5), pre-injection hypocalcemia, male sex, hypophosphatemia, hypomagnesemia, and vitamin D insufficiency.
The prevalence of hypocalcemia was higher in our study (6%) than in the original clinical trial (FREEDOM), which reported a prevalence of 0–0.1% [
4,
6]. A low prevalence of hypocalcemia (< 1% hypocalcemia) was also reported in the FREEDOM sub-analysis of patients with renal failure and in a post-marketing real-world prospective observational trial [
12,
17]. Our findings are in the same range as other large-scale studies on real-world cohorts, which reported the frequency of denosumab-induced hypocalcemia to range from 4 to 8% [
7,
8,
10].
Previous hypocalcemia data in large cohorts, including clinical trials, was mainly based on calcium analysis before consecutive injections (e.g., every 6 months) or other predefined intervals not consequently taken shortly after injection [
6‐
8,
12]. As post-injection hypocalcemia is mostly seen in the weeks following injection [
9,
13,
14], there is a risk of underestimation of the prevalence of hypocalcemia if longer intervals between injection and calcium analysis are allowed. Another explanation for the difference is that patients are more systematically controlled in clinical trials, and possible insufficiencies of calcium and vitamin D will either exclude the patient from the study or delay inclusion until adequately supplemented. Furthermore, patients included in a clinical trial are probably more adherent to supplements, e.g., calcium and vitamin D supplements, which might also affect the lower hypocalcemia frequency observed in these studies. Additionally, previous recommendations, also used in the FREEDOM trial, were 1000 mg calcium supplementation daily, which is twice as high as our national recommendations, which might have affected the results in patients with borderline calcium sufficiency. Accordingly, previous studies highlight the importance of adequate calcium supplements to prevent denosumab-induced hypocalcemia [
7,
13]. This could raise the question of whether the general recommendation of 500 mg calcium/day is enough. Whether a higher dose should be recommended to patients on denosumab or preferentially to patients with certain risk profiles remains to be determined.
Using non-routine data, i.e., calcium analysis only if doctors ask for it [
17], might include selection bias where only high-risk patients are followed up with calcium measurements. This might yield an overestimation of the prevalence of hypocalcemia, or it might risk missing cases of mild hypocalcemia.
Most previous studies analyzed total calcium, which includes not only the biologically active form (ionized calcium) but also albumin bound and other calcium complexes [
11]. In contrast, we analyzed ionized calcium directly, resulting in the assessment of biologically active calcium being more precise [
11,
18] and the rate of post-injection hypocalcemia more reliable.
Renal failure is a known risk factor for post-injection hypocalcemia [
7,
8,
13,
19], which is in line with our results of a clearly elevated risk of hypocalcemia in patients with severe renal failure, i.e., CKD 4 and 5 (GFR < 30 [mL/min]/1.73 m
2), and a tendency through non-significant in patients with CKD 3b (GFR 30–44 [mL/min]/1.73 m
2). In contrast, CKD 3a (GFR 45–60 [mL/min]/1.73 m
2) did not increase risk of hypocalcemia in our study. In renal failure, conversion of 25-OH vitamin D to the active form calcitriol is impaired, rendering a risk of negative calcium balance where calcium uptake from the intestine is insufficient and the body is dependent on the bone for maintaining normal blood calcium levels. Clinically this is seen by secondary hyperparathyroidism with low or normal (compensated) blood calcium values. In this scenario, an effective antiresorptive agent, such as denosumab, blocks the necessary efflux from bone, resulting in hypocalcemia. To decrease this risk, it is important to adjust the calcium balance before treatment, e.g., with calcitriol, as well as to ascertain adequate calcium supplements.
In our study we found a higher risk of post-injection hypocalcemia in men than in women. Similar findings have been reported previously [
7,
8]; however, in a post-analysis of previous data, it was concluded that this was more likely secondary to selection bias [
20]. Despite correcting for renal failure and age in our study, the gender difference remained. Whether this is an actual gender difference or cofactors that were not measured and adjusted for in our study is unclear.
Pre-injection hypocalcemia has previously been reported as a risk factor for hypocalcemia [
7,
8]. However, in our cohort, pre-injection hypocalcemia was not significantly associated with severe cases of hypocalcemia, and most patients of post-injection hypocalcemia had normal pre-injection calcium levels, thus weakening pre-injection hypocalcemia as a single clinical risk marker to identify high-risk patients (sensitivity: 9%).
In our study, we included measurements of magnesium and phosphate; both minerals are tightly connected to the calcium balance [
21,
22]. Low pre-injection values of magnesium or phosphate imposed a substantially increased risk of post-injection hypocalcemia. This could reflect signs of malnutritional status in patients with hypocalcemia, where calcium levels are dependent on calcium from bone. Some of the patients had intestinal diseases with a risk of malabsorption as well as individuals with a clinical suspicion of malnutrition. In addition, one patient had a phosphate-associated disease. Both magnesium and phosphate minerals are easy and inexpensive to analyze, and the present study indicates that they might be valuable markers to predict the risk of post-injection hypocalcemia. However, in the present study, these minerals were not routinely evaluated in all patients, and, especially regarding phosphate, there were few observations (5% of injections) available. Thus, it would be valuable to confirm the findings in a larger cohort.
Post-injection hypocalcemia not only occurred after the first injection, the prevalence was similar throughout the treatment period (3–8%). This is in concordance with the study by Tsvetov et al. [
8]. The finding of unchanged prevalence rates seems feasible as risk factors, e.g., renal failure progression and malnutrition, might develop over time as patients age. On the other hand, most patients with post-injection hypocalcemia did not have recurrent hypocalcemia, which was possibly caused by a more alert preventive approach on the following injections, e.g., use of extra calcium and vitamin D as well as other preventive treatments such as nutrition optimization.
Severe hypocalcemia was rare, i.e., following 1% of all injections. In five cases, patients were admitted to the hospital or required an acute outpatient visit after the injection. These patients had a complicated disease history, including severe kidney disease, intestinal disease, and/or malnutrition.
Measuring post-injection laboratory values are costly, including analysis costs and organizational costs with staff dedicated to organizing testing and follow-up of the results. Furthermore, the extra follow-up causes inconvenience for the patients and might involve relatives/personal to assist as there will be an extra visit to the laboratory and phone contact with nurse. Therefore, although the risk of hypocalcemia needs to be addressed and high-risk individuals, as described above, need to be identified, the vast majority (94%) of injections were not followed by hypocalcemia. Thus, post-injection calcium screening does not seem motivated in general.
Furthermore, the clinical significance of mild and moderate post-injection hypocalcemia is of uncertain clinical significance. Some cases of mild hypocalcemia may even reflect a normal biological variance, i.e., when there is a small decrease in calcium from the normal reference. We do not have a least significant change (LSC) value or reference change value (RCV) for ionized calcium. Assuming that it behaves in a similar manner to total calcium, we might expect an RCV of approximately 6%. When applying this to our data, 21 cases of mild hypocalcemia could possibly reflect normal variation. Excluding these from the analysis would yield a total post-injection hypocalcemia frequency of 4.5% instead of 6.3% (rates of moderate and severe hypocalcemia are not altered). However, this assumes similar behavior in the two calcium analysis methods.
Similar to previous reports, we believe that patients with moderate and severe renal failure need to be followed up more carefully, possibly including routine post-injection calcium measurements. Other risk groups identified are patients with malnutrition/malabsorption. In both risk groups, adequate pretreatment is important to ascertain a balanced calcium system that is not dependent on calcium from bone. Thus, it is necessary to administer adequate calcium and vitamin D3 supplementation as well as in renal failure caltiriol treatment if required. Chronic magnesium insufficiency may blunt the PTH response and thus disable the compensation mechanism in hypocalcemia, which is why adequate magnesium is important. Further studies on the value of including magnesium, phosphate, 25-OH vitamin D, and PTH in the pre-testing would be valuable.
Strengths and limitations
In our study, pre- and post-injection calcium values were available for all injection time points, whereas the other laboratory values were taken less frequently (e.g., creatinine GFR, which was recommended once every 12 months, and magnesium, often before the first injection) or non-routinely (e.g., phosphate, 25-OH-vitamin D). Therefore, there is a risk of both over- and underestimating the impact of these factors. Phosphate was only taken in 5% of cases, including one patient with a known phosphate disturbance, which could affect the result. We chose to include all patients as we believe that this best mirrors a real-world cohort. The low and non-systematically measured values of vitamin D and PTH is a clear limitation and risks reducing the power of the data as well as creating selection bias. Other limitations of this study include the lack of data on symptoms and dosing of calcium and vitamin D supplements at each injection time point. The former was not systematically included in the case records, and the latter could not be extracted retrospectively from the case records.
The strengths of the study are the study design, including a typical real-world osteoporosis cohort, which results in the data being generalizable. In addition, the study design, with routinely analyzed post-injection calcium measurements in a well-defined time-period between the injection and follow-up value (i.e., maximum of 31 days), thus avoiding selection bias, is a strength.
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