Preventive Role of Vitamin D Supplementation for Acute Phase Reaction after Bisphosphonate Infusion in Paget’s Disease

Abstract

Context

Intravenous aminobisphosphonates (N-BPs) can induce an acute phase reaction (APR) in up to 40% to 70% of first infusions, causing discomfort and often requiring intervention with analgesics or antipyretics.

Objective

Our aim was to explore the risk factors of APR in a large sample of patients with Paget’s disease of bone (PDB) and to assess the possible preventive effects of vitamin D administration.

Methods

An observational analysis was performed in 330 patients with PDB at the time of N-BP infusion. Then, an interventional study was performed in 66 patients with active, untreated PDB to evaluate if vitamin D administration (oral cholecalciferol 50 000 IU/weekly for 8 weeks before infusion) may prevent APR.

Results

In a retrospective study, APR occurred in 47.6% and 18.3% of naive or previously treated patients, respectively. Its prevalence progressively increased in relation to the severity of vitamin D deficiency, reaching 80.0% in patients with 25-hydroxyvitamin D (25OHD) levels below 10 ng/mL (relative risk (RR) = 3.7; 95% confidence interval (CI) 2.8–4.7, P < .0001), even in cases previously treated with N-BPs. Moreover, APR occurred more frequently in patients who experienced a previous APR (RR = 2.8; 95% CI 1.5–5.2; P < .001) or in carriers of SQSTM1 mutation (RR = 2.3; 95% CI 1.3–4.2; P = .005). In the interventional study, vitamin D supplementation prevented APR in most cases, equivalent to a RR of 0.31 (95% CI 0.14–0.67; P < .005) with respect to prevalence rates of the observational cohort. A similar trend was observed concerning the occurrence of hypocalcemia.

Conclusions

The achievement of adequate 25OHD levels is recommended before N-BP infusion in order to minimize the risk of APR or hypocalcemia in PDB.

Paget’s disease of bone, acute phase reaction, bisphosphonates, vitamin D, parathyroid hormone, bone turnover

Paget’s disease of bone (PDB) is a chronic skeletal disorder affecting up to 1% to 5% of the elderly population in certain European countries, which typically results in enlarged and deformed bones in one or more regions of the skeleton (1–4). Specific therapy for PDB is aimed at decreasing the abnormal bone remodeling, and bisphosphonates (BPs) are currently considered the treatment of choice (5, 6). These compounds are associated with a reduction in bone turnover markers and an improvement in radiological and scintigraphic appearance as well as with a reduction in bone pain, whereas the effects of treatment on the development or progression of PDB complications remain poorly understood (5–7). To date, different BPs have been successfully used for the treatment of PDB. In particular, the development of newer, more potent nitrogen-containing BPs (N-BPs) has remarkably improved treatment outcomes (7). These compounds have a high binding affinity to hydroxyapatite as well as increased potency in terms of inhibition of bone resorption (8–10), and can be given as either oral (alendronate and risedronate) or intravenous (zoledronate and neridronate) regimens.

All N-BPs administered intravenously can induce an acute phase reaction (APR), essentially characterized by pyrexia, musculoskeletal pain, and other flu-like symptoms, which may require intervention with analgesics and antipyretics (11–14). These symptoms are transient and occur predominantly on first exposure to the drug in most N-BP-“naïve” patients (up to 40–70% of cases) and rarely with subsequent doses (15). However, despite its higher prevalence, preventative strategies against APR are not well established, so that APR remains the main reason leading to treatment discontinuation in N-BP users (16).

Although APR was first described in 1987 (17), the underlying molecular mechanism has only recently become clearer. Initial reports demonstrated that the flu-like symptoms are generally accompanied by decreased lymphocyte counts and elevated levels of the proinflammatory cytokines such as interleukin (IL-6), interferon (IFN)-γ, and tumor necrosis factor (TNF)-α (18, 19). Moreover, this adverse event seems to be related to the accumulation of isoprenyl pyrophosphate (IPP), the metabolite immediately upstream of farnesyl pyrophosphate (FPP) synthase in the mevalonate pathway, by cells in peripheral blood (most likely monocytes) due to the inhibition of FPP synthase by N-BPs. IPP is known to be a ligand for the most common subset of γδ-T cells in humans, Vγ9Vδ2 T cells (20). Although the precise mechanism by which IPP is released or “presented” to these γδ-T cells remains unknown, their activation perhaps via a selective receptor, causes the release of TNF-α, IL-6, and IFN-γ and thereby initiates the proinflammatory APR. In support of this hypothesis, Vγ9Vδ2⁺ T cells have been shown to recognize endogenous mevalonate metabolites, such as IPP produced by zoledronate-pulsed tumor cells (21). Independently, it was also demonstrated that the ability of N-BPs to activate Vγ9Vδ2⁺ T cells closely matches their ability to inhibit FPP synthase (22, 23), and that cotreatment with a statin (to prevent the accumulation of IPP) completely abrogates the stimulatory potential of N-BPs on γδ T cell activation and proliferation in vitro (22, 24). Despite the latter observation raising the intriguing possibility that statins could be used to prevent APR, this was not demonstrated by placebo-controlled studies on postmenopausal women with osteoporosis (25–27).

More recently it has been shown that both γδ T cells and monocytes become rapidly activated after treatment with zoledronate and that the proportion of circulating γδ T cells before infusion is an important determinant of the occurrence of APR (28, 29). At the same time, both intravenous and oral N-BP regimens may decrease circulating γδ T cells for at least 1 year (30), making a likely explanation for the reduced prevalence of APR with subsequent N-BP infusions.

Of interest, a previous observation in 90 postmenopausal women treated with N-BPs for osteoporosis evidenced a negative association between circulating 25 hydroxyvitamin D (25OHD) levels and the development of APR, likely due to immunomodulatory effects of vitamin D on γδ T cells (31). However, this association remains to be demonstrated in larger samples and in patients with PDB, where the levels of proinflammatory cytokines such as IL-6 are per se increased (32, 33). In our previous studies in PDB patients treated with either intravenous (zoledronate or neridronate) or intramuscular (neridronate) N-BPs APR occurred in about 15% to 30% of patients and mostly in previously untreated cases (34, 35). Indeed, in a preliminary analysis of 56 PDB patients treated with neridronate those experiencing an APR showed lower vitamin D levels before treatment than those without the APR, either before or after the exclusion of subjects with previous N-BP treatment (35).

Thus, the aim of our study was to clarify the characteristics and the risk factors of APR through a retrospective analysis of a large sample of pagetic patients after N-BP treatment and then to prospectively assess the effect of preventive vitamin D administration on the prevalence of APR in 66 patients with 25OHD levels below 20 ng/mL (50 nmol/L).

Subjects and Methods

Patients and study design

This study involved the following 2 cohorts of PDB patients.

Observational cohort.

We firstly performed a retrospective analysis of 330 consecutive patients with active PDB who had been previously treated with neridronate 200 mg intravenously (IV) (100 mg on 2 consecutive days, n = 142), or zoledronate 5 mg IV (n = 188). Of these patients, 128 (38. 8%) were naïve from any treatment at the time of infusion, while 202 (61.2%) had already been treated with other BPs (clodronate 4.5%, pamidronate 42.1%, risedronate 11.4%, neridronate 36.6%, zoledronate 5.4%). All included patients had follow-up information for at least 12 months from treatment, with detailed data about adverse events occurring after treatment, including APR. Adverse events were categorized according to the individual preferred terms indicated in the Medical Dictionary for Regulatory Activities (15, 36).

Prospective cohort.

Then, we performed a prospective study in 66 consecutive PDB cases, at the time of diagnosis and thus naïve for any previous BP treatment, that were treated with neridronate 200 mg IV (100 mg in 2 consecutive days, n = 35) or zoledronate 5 mg IV (n = 31). Eligibility was defined by the presence of serum total alkaline phosphatase (ALP) above the upper limit of the normal range (120 UI/L) on 2 consecutive measurements and/or the presence of bone pain at affected sites. Exclusion criteria included major comorbidity, metabolic bone disease other than PDB, recent fracture at pagetic bone, clinically significant liver disease, and renal impairment. In all patients the diagnosis of PDB was confirmed by bone scintigraphy and X-ray examination of areas of increased isotope uptake, unless recently performed. At the time of diagnosis, all patients had baseline 25OHD measurement. In the presence of 25OHD values below 20 ng/mL, patients were advised to receive cholecalciferol 50 000 IU/weekly for 8 weeks, before N-BP infusion, according to Holick and colleagues (37, 38). We decided for ethical reasons (due to the observed increased risk of APR and hypocalcemia) to give the vitamin D replacement regimen to all patients with baseline 25OHD < 20 ng/mL and not to include a placebo arm. After N-BP infusion, all patients were advised to receive calcium plus vitamin D supplementation throughout the study period (1 g of calcium and 800 IU cholecalciferol per day). The occurrence of APR was recorded at baseline and after 24 hours, 48 hours, 7 days, and 10 days from N-BP infusion by a phone call evaluating the presence of pyrexia and/or myalgia, the duration and the severity of symptoms (mild, moderate, severe), and the use of analgesics, paracetamol, or other anti-inflammatory drugs. All other adverse events were recorded during follow-up visits after 3 days and at 1, 6, and 12 months from infusion, and their severity and relationship to treatment were evaluated.

The study was approved by the local Ethics Committee and written informed consent was obtained from all participants. General characteristic of both observational and interventional cohorts are shown in Table 1.

Table 1.

General characteristics of PDB cases in both observational and interventional cohorts.

	Observational cohort (n = 330)	Interventional cohort (n = 66)
Age (yr)	75.1 ± 11.2	73.8 ± 12.4
Age at diagnosis (yr)	58.3 ± 12.7	56.5 ± 11.8
Sex (M/F)	208/122	39/27
Monostotic PDB cases	135 (40.9%)	20 (31.2%)
Polyostotic PDB cases	195 (59.1%)	46 (68.8%)
Affected skeletal sites (n)	2.6 ± 2.2	2.3 ± 1.3
BMI	26.9 ± 4.1	26.4 ± 3.5
Total ALP (IU/L)	284 ± 82	276 ± 77
Baseline 25-OH vitamin D (ng/mL)	23.4 ± 11.3	24.2 ± 13.5
25-OH vitamin D < 30 ng/mL	261 (79.1%)	53 (80.3%)
Prevalence of SQSTM1 mutation	45 (13.6%)	8 (12.1%)

Biochemical evaluation

All subjects in the observational and interventional cohorts, respectively, had been or were evaluated at baseline, and after 1, 6, and 12 months from treatment. At each time point, venous blood was collected in the fasting state for serum analysis. Moreover, an additional blood sampling was planned at day 3 from infusion in the interventional cohort.

Serum ALP, aspartate and alanine aminotransferases, ionized and total calcium (Ca), phosphate (P), and creatinine levels were determined by standard methods. Bone ALP (bALP, Alkphase-B, Metra Biosystem, Mountain View, CA), serum C-terminal telopeptides of Type I collagen (sCTX, serum CrossLaps; Osteometer, Herlev, Denmark), 25OHD (LIASON 25-OH Vitamin D TOTAL Assay; Diasorin Diagnostics), and intact parathyroid hormone (PTH; N-tact PTH, IRMA kit, DiaSorin, Stillwater, MN) were also evaluated. The observed intra- and interassay coefficients of variation for each marker were, respectively, as follows: below 1.6% for ALP, 2.0% and 4.1% for bALP, 6.1% and 5.4% for sCTX, 2.7% and 10.9% for 25OHD, and 2.8% and 4.0% for PTH. All analyses were performed in the clinical and research laboratory of the Department of Medicine Surgery and Neurosciences, University of Siena. The quality and accuracy of the 25OHD and PTH analyses from our laboratory are validated on an ongoing basis by participation in the External Quality Evaluation Program from the “Centro di Riferimento per la Qualità dei Servizi di Medicina di Laboratorio” (39). Finally, serum levels of IL-6 were determined from stored blood samples collected at the time of infusion and after 3 days, 1 month, and 6 months in all the PDB cases from the interventional cohort using a commercially available ELISA kit (Quantikine, R&D Systems, Minneapolis, USA, intra- and interassay coefficient of variation (CV) < 5%).

Statistical analysis

The proportion of patients who developed an APR was compared between treatment groups using the standard chi-square or Fisher exact test. Biochemical data for each treatment group and interactions between treatment and time or vitamin D status were evaluated by analysis of variance and covariance, adjusted for age and body mass index (BMI). Linear regression analysis with Pearson’s correlation coefficient was used to determine the relationships between biochemical parameters. Relative risk (RR) and 95% confidence interval (CI) were calculated to assess the association between 25OHD deficiency or other categorical variables and APR. Analysis was performed using Statistica 5.1 (Statsoft Inc., Tulsa, OK) and SPSS (release 6.1, Chicago, IL).

Results

Observational study

The mean age of patients was 75 years (range 43–98 years) with a mean age at diagnosis of 58 years; 40.9% of them were affected by a monostotic form of PDB, while 59.1% had a polyostotic disease. All patients had been previously screened for SQSTM1 mutation, with 4 mutations (P392L, M404V, Y383X, E396X) identified in 45/330 (13.6%) of cases.

Overall, both N-BP treatment regimens were well tolerated. Major adverse events for each treatment group occurring within 12 months from infusion are summarized in Table 2. These were mainly related to APR with influenza-like symptoms, such as pyrexia and myalgia, in 98/330 (29.7%) of cases: 67/219 (30.6%) in the zoledronate group and 31/111 (27.9%) in the neridronate group. As shown in Table 3, these symptoms were significantly more frequent in treatment-naïve patients than in previously treated patients (47.6% vs. 18.3%, P < .0001), equivalent to a RR of 2.6 (95% CI 1.8–3.7; P < .0001). All APR symptoms were mild to moderate in severity and generally resolved within few days (mean duration 2.7 ± 2.0 days) with (66.3%) or without (33.5%) the use of oral paracetamol or other anti-inflammatory drugs.

Table 2.

Adverse events following the last amino-bisphosphonate treatment in observational retrospective study (n = 330).

	Zoledronate IV (n = 219)	Neridronate IV (n = 111)
Acute phase reaction, n (%)	67 (30.6)	31 (27.9)
Influenza-like illness	40 (18.3)	29 (26.1)
Pyrexia	38 (17.3)	28 (25.2)
Myalgia	40 (18.3)	29 (26.1)
Fatigue	18 (8.2)	13 (11.7)
Headache	15 (6.8)	10 (9.0)
Diarrhea, n (%)	5 (2.3)	3 (2.7)
Hypocalcemia, n (%)	13 (11.7)	10 (9.0)
Dermatitis, n (%)	1 (0.46%)	0
Atrial fibrillation, n (%)	0	0
Osteonecrosis of the jaw	0	0

Table 3.

General characteristics of PDB cases experiencing or not APR after intravenous bisphosphonate treatment in the observational retrospectivecohort.

	PDB cases experiencing APR (n = 98)	PDB cases not experiencing APR (n = 232)
Age, years	72.05 ± 13.2	73.57 ± 11.3
Age at diagnosis, years	58.1 ± 10.7	58.6 ± 11.5
Sex (M/F), n	48/33	147/102
Affected skeletal sites (n)	2.62 ± 1.13	2.51 ± 1.08
Zoledronate treated pts, n = 219, n (%)	67 (30.6)	152 (65.1)
Neridronate treated pts, n = 111, n (%)	31 (27.9)	80 (72.1)
Naïve patients, n = 128, n (%)	61 (47.6)a	67 (52.3)
Previous treated patients, n = 202, n (%)	37 (18.3)a	165 (81.7)
Use of NSAID, n (%)	18 (18.4)	57 (24.6)
Use of statins, n (%)	15 (15.3)	54 (23.3)

^a  P < 0.0001 naïve vs previous treated patient.

Neither gender nor disease severity nor use of other drugs except previous N-BP treatment was significantly associated with the occurrence of APR (Table 3). However, we observed a trend for a reduced incidence of APR in statins users (15.3% vs. 23.3% in users and nonusers, respectively, P = .09). Overall, 126 PDB cases were treated for PDB due to biochemical relapse (ALP levels above the normal reference range), 72 due to the referral of symptoms (mostly pain) in the presence of normal ALP levels, while the remaining 132 cases had biochemical relapse in association with symptoms. The prevalence of APR was significantly higher (P < .01) in those cases with biochemical relapse either in the presence (31.8%) or the absence of symptoms (31.7%), than in patients with normal ALP levels at time of infusion (22.2%).

Within the group of patients previously treated with BPs, the prevalence of APR was inversely related with the antiresorptive potency of the previous BP regimen, occurring, respectively, in 44.4%, 23.5%, 17.3%, 10.8%, and 9.1% of clodronate-, pamidronate-, risedronate-, neridronate-, and zoledronate-treated cases. Moreover, APR prevalence rates also differed in relation to the number of previous BP treatment courses, being less prevalent in patients with 3 or more courses (12.6%) than in those with 1 (24.6%) or 2 (21.4%) courses (P = .05), while they were not significantly influenced by the time from the previous treatment course. Of interest, among previous BP users, APR occurred more frequently in those patients who had experienced a previous APR than in the remaining cases (30.0% vs. 10.6%, respectively, P < .005), with a RR of 2.8 (95% CI 1.5–5.2; P < .001), as well as in carriers of SQSTM1 mutation (35.5% vs. 15.2%, P < .01), with a RR of 2.3 (95% CI 1.3–4.2; P = .005).

Measurements of 25OHD levels were available for all patients at the time of treatment with intravenous neridronate or zoledronate. The detailed characteristics and the prevalence of PDB patients with “vitamin D insufficiency” (25OHD levels between 20 and 30 ng/mL equivalent to 50 and 75 nmol/L, respectively), “vitamin D deficiency” (25OHD below 20 ng/mL or 50 nmol/L), and “severe hypovitaminosis D” (25OHD below 10 ng/mL or 25 nmol/L) are shown in Table 4. Overall, we observed a remarkable prevalence of hypovitaminosis D (79.1%), defined as 25OH Vitamin D levels below 30 ng/mL, in our PDB patients with only 20.9% of cases showing “normal” 25OHD levels. Even when using the more conservative threshold of 20 ng/mL, as proposed by the Institute of Medicine (40), 41.2% of patients resulted deficient in vitamin D.

Table 4.

Vitamin D status in observational retrospective study (n = 330).

	Mean 25OHD (ng/mL)	25OHD > 30 ng/mLn (%)	25OHD < 30 ng/mLn (%)	25OHD 30-20 ng/mLn (%)	25OHD < 20 ng/mLn (%)	25OHD < 10 ng/mLn (%)
Total population	23.42 ± 11.32	69 (20.9)	261 (79.1)	125 (37.9)	136 (41.2)	45 (13.6)
Men	23.40 ± 9.90	41 (12.4)	167 (50.6)	80 (24.2)	87 (26.4)	22 (6.7)
Women	23.44 ± 13.45	28 (8.5)	94 (28.5)	45 (13.6)	49 (14.8)	23 (7.0)
Monostotic PDB	23.36 ± 10.86	28 (8.5)	107 (32.4)	51 (15.4)	56 (17.0)	20 (6.10)
Polyostotic PDB	23.65 ± 11.62	41 (12.4)	154 (46.7)	74 (22.4)	80 (24.2)	25 (7.6)
BMI < 18.5	12.72 ± 9.40	0 (0)	4 (1.2)	1 (0.3)	3 (0.9)	2 (0.6)
BMI 18.5–24.99	24.05 ± 11.82	26 (7.9)	68 (20.6)	31 (9.4)	37 (11.2)	15 (4.5)
BMI 25–30	23.54 ± 11.30	33 (10.0)	123 (37.3)	64 (19.4)	59 (17.9)	19 (5.7)
BMI > 30	22.57 ± 9.10	10 (3.0)	66 (20.0)	29 (8.0)	37 (11.2)	9 (2.7)

As it is evident in Table 4, there were no significant differences in 25OHD levels in relation to the gender or the skeletal extent of PDB. Consistent with previous findings in the general population, we found a significant correlation between vitamin D and BMI subclasses (P < 0.05), with significantly reduced 25OHD levels in underweight and obese patients with respect to patients with normal weight (Table 4).

At the time of intravenous N-BP infusion, 25OHD levels were lower in treatment-naïve patients with respect to PDB cases who had been previously treated with other BPs (mean values 19.7 ± 8 ng/mL vs. 25.8 ± 12 ng/mL, respectively, P < .0001). This was likely due to the higher proportion of patients taking vitamin D supplements in the previously treated group than in treatment-naïve patients (63.9% vs. 22.4%, respectively, P < .0001). Of interest, 25OHD levels were lower in PDB cases experiencing APR than in patients who did not had APR after N-BP infusion, either before (16.0 ± 9 ng/mL vs. 26.6 ± 11 ng/mL; P = .005) or after the exclusion of subjects with a previous N-BP treatment (16.5 ± 9 ng/mL vs. 22.8 ± 8 ng/mL; P < .005). Likewise, significantly lower 25OHD levels were observed in patients experiencing APR when previously treated patients, patients with or without ongoing vitamin D replacement, or patients with normal or higher BMI were considered separately. Indeed, within the subgroup of patients reporting ongoing vitamin D supplementation only 9/122 (7.3%) cases with 25OHD levels above 20 ng/mL at the time of infusion had APR, versus 12/38 (31.6%) cases with APR reported in those patients with 25OHD below 20 ng/mL (P = .0001). Remarkably, in the overall group, the prevalence of APR progressively increased in relation to the severity of vitamin D deficiency, reaching 80.0% (36/45) in patients with 25OHD levels below 10 ng/mL. Accordingly, the RRs of APR were respectively 3.7 (95% CI 2.8–4.7, P < .0001), 4.2 (95% CI 2.8–6.2, P < .0001), and 2.3 (95% CI 1.3–4.2, P < .01) in patients with 25OHD below 10 ng/mL, 20 ng/mL, and 30 ng/mL.

Such a higher prevalence of APR in patients with severe vitamin D deficiency was maintained not only in treatment-naïve cases (20/25, 80.0%) but also in PDB cases previously treated with N-BPs (16/20, 80.0%) (Fig. 1). Moreover, patients with APR showed enhanced bone turnover at the time of infusion compared with those without APR, as documented by increased levels of bALP (55.6 ± 49.3 μg/L vs. 42.6 ± 42.7 μg/L; P = .01) and sCTX (1.198 ± 1.604 ng/mL vs. 0.928 ± 0.610 ng/mL; P = .01). This difference became not significant when only treatment-naïve patients or polyostotic cases were considered.

Figure 1.

Prevalence of acute phase reaction according to 25OH vitamin D levels in naive and previous treated pagetic patients. Prevalence rates were highest reaching 80% of cases in the presence of severe hypovitaminosis D (25OHD < 10 ng/dL), irrespective of previous aminobisphosphonate treatment.

Calcium levels at baseline were in the normal range in most patients, with only 6 cases (1.8%) of mild hypocalcemia (serum Ca between 8.0 and 8.4 mg/dL). After infusion 23 patients (6.9%) showed hypocalcemia, 3 of them had severe hypocalcemia (serum Ca < 8 mg/dL) and 20 had mild hypocalcemia. In these patients experiencing hypocalcemia after N-BP, 25OHD levels were significantly reduced than in patients without hypocalcemic events (Fig. 2). Likewise, this group of patients with hypocalcemia also showed a statistically significant increase in circulating PTH (50.5 ± 16.7 vs. 40.4 ± 13.9 pg/mL; P < .05) and a reduction in serum P (2.86 ± 0.4 vs. 3.28 ± 0.5 mg/dL; P < .01) with respect to the rest of the cohort with normal calcium levels after infusion. No cases of atrial fibrillation or osteonecrosis of the jaw were reported at 6, 12, or more months from N-BPs infusion (with a duration of observation ranging from 12 to 24 months from infusion).

Figure 2.

25OH vitamin D levels in pagetic patients who experienced or not hypocalcemic events after aminobisphosphonate infusion.

Biochemical analyses from 1 to 12 months from infusion evidenced an increase in PTH levels at 1 month after N-BP infusion, along with a decrease in 25OHD and P levels (Table 5). These differences were more pronounced in patients with APR than in those without APR. Indeed, while baseline Ca levels did not significantly differ between cases with or without APR, P levels were lower and PTH levels were higher in the APR group. Accordingly, the prevalence of secondary hyperparathyroidism at baseline was 13/98 (13.3%) vs. 14/232 (6.0%) in the APR and non-APR groups, respectively (P < .05). Conversely, the prevalence rate of hypophosphatemia at baseline was low and did not significantly differ between patients that developed or not APR (2.3% vs. 2.6%, respectively). In particular, just one case showed serum P levels below 2.0 mg/dL. Moreover, patients experiencing APR also showed higher bALP and sCTX at different time points from N-BP treatment than PDB cases without APR (Table 5).

Table 5.

Variation of biochemical markers after intravenous N-BP treatment in the observational retrospective study in PDB cases with or without APR.

	Baseline	1 month	6 months	12 months
Ca (mg/dL)
APR	9.2 ± 0.6	9.2 ± 0.6	9.4 ± 0.5	9.5 ± 0.5
No APR	9.3 ± 0.5	9.3 ± 0.4	9.4 ± 0.5	9.5 ± 0.4
P (mg/dL)
APR	3.2 ± 0.6 c	3.0 ± 0.6 c	3.2 ± 0.6	3.3 ± 0.5
No APR	3.4 ± 0.5	3.2 ± 0.5	3.3 ± 0.5	3.4 ± 0.5
25OHD (ng/mL)
APR	16.0 ± 9d	21.7 ± 10d	26.1 ± 11d	25.2 ± 10 d
No APR	26.6 ± 11	26.6 ± 10	31.3 ± 13	30.6 ± 11
PTH (pg/mL)
APR	45.3 ± 17b	65.6 ± 30d	44.5 ± 18a	45.9 ± 20
No APR	39.2 ± 16	51.5 ± 20	40.1 ± 16	42.7 ± 18
BALP (µg/L)
APR	54.7 ± 37d	46.6 ± 36d	16.0 ± 12d	16.4 ± 15 c
No APR	38.9 ± 30	23.9 ± 14	12.1 ± 6	12.3 ± 7
CTX (ng/mL)
APR	1.18 ± 1.0 d	1.59 ± 3.2 a	0.51 ± 0.6	0.46 ± 0.5
No APR	0.85 ± 0.5	0.75 ± 2.1	0.44 ± 0.4	0.38 ± 0.3

^a  P < 0.05; ^bP < 0.01; ^cP < 0.005; ^dP < 0.001

Interventional study

Based on the results from the observational analysis, we then performed a prospective, interventional, study in 66 patients at the time of PDB diagnosis (Table 1). As is evident, and consistent with the results from the observational study, hypovitaminosis D was common also in this cohort of patients (mean 25OHD 24.2 ± 13 ng/mL), with 43 cases (65.1%) below the 20 ng/mL threshold. According to the established protocol, all the 43 patients with 25OHD levels below 20 ng/mL received oral vitamin D3 50 000 IU/weekly for 8 weeks before N-BPs infusion with either zoledronate 5 mg (n = 21/30) or neridronate 200 mg (n = 22/36). At the time of N-BP infusion mean 25OHD levels were 42.6 ± 14.2 ng/mL in PDB cases without vitamin D deficiency at recruitment (25OHD > 20 ng/mL) while significantly raised to 34.3 ± 5.4 ng/mL (P < .0001 vs. baseline) in the group of 43 PDB patients with vitamin D deficiency (25OHD < 20 ng/mL) at the time of diagnosis, who had received vitamin D supplementation for 8 weeks before infusion.

All adverse events occurring within the first 10 days and after 1, 6, and 12 months from N-BP infusion were recorded. Of interest APR was less common, and occurred in only 6/66 patients (9.1%), with a statistically significant reduced prevalence than that described in the overall cohort of patients from the observational study, as well as in naïve cases from the observational cohort (Fig. 3). This was equivalent to a RR of 0.31 (95% CI 0.14–0.67; P < .005) with respect to prevalence rates in the overall observational cohort and of 0.19 (95% CI 0.09–0.42; P < .0001) with respect to observational cases naïve to N-BPs. The characteristics of APR as pyrexia and myalgia were milder in the interventional cohort with respect to what observed in the observational study. Moreover, no cases of mild or severe hypocalcemia, atrial fibrillation, or other long-term complications were reported within the first 12 months from N-BP infusion. A higher but not significant prevalence of APR was observed in zoledronate (4/30, 13.3%) than in neridronate (2/36, 5.5%) treated patients. There were no significant differences in 25OHD levels at the time of N-BP infusion between patients who developed or not APR (34.5 ± 3.9 ng/mL, vs. 37.5 ± 10.5 ng/mL, respectively). However, all the 6 cases of APR occurred in patients with baseline 25OHD < 20 ng/mL, who received vitamin D supplementation for 8 weeks before infusion, and 4 of them had SQSTM1 mutation. In 5 of these cases 25OHD levels at the time of infusion were above the “normal” range of 30 ng/mL. We did not observe any significant difference in APR severity according to baseline 25OHD levels or SQSTM1 mutations.

Figure 3.

Prevalence rates of acute phase reaction in observational and interventional studies. Vitamin D supplementation in the interventional study significantly reduced the occurrence of APR in aminobisphosphonate naive patients to less than 10%, as compared to 45%, as observed in aminobisphosphonate naive cases from the observational study.

Consistent with the results of the observational study, serum PTH increased significantly at 3 days and 1 month from infusion (Fig. 4). This increase was higher in patients with APR than in those without APR and was associated with a parallel decrease in 25OHD, Ca and P levels. Moreover, patients with APR also showed significantly higher bALP and sCTX levels between 6 and 12 months from treatment than patients without APR.

Figure 4.

Serum levels of 25OH vitamin D, PTH, Ca, P, sCTX and bALP after aminobisphosphonate infusion in pagetic patients from the interventional study who experienced or not an acute phase reaction. A remarkably increased peak of PTH was observed after aminobisphosphonate infusion in those cases experiencing an acute phase reaction.

At the time of infusion, IL-6 levels did not differ between patients that developed or not an APR and increased significantly at 3 days and 1 month post infusion in most patients (Fig. 5). Of interest the increase in IL-6 was more pronounced in patients developing APR, with a statistically significant difference at 3 days and 1 month from infusion with respect to patients that did not develop APR. Moreover a strong and positive correlation was observed between PTH and IL-6 levels at 3 days (r = 0.65; P < .0001), 1 month (r = 0.48; P < .001) and 6 months (r = 0.31; P < .05) from infusion.

Figure 5.

Serum IL-6 levels before and after aminobisphosphonate infusion in pagetic patients from the Interventional study who experienced or not an acute phase reaction.

Discussion

Vitamin D deficiency is a common issue in elderly people, with the inclusion of subjects treated with BPs for osteoporosis. Also in our study, we observed a remarkable prevalence of hypovitaminosis D in patients with PDB either considering the 30 ng/mL (79%) or the 20 ng/mL (41%) threshold. The deficit in 25OHD was more pronounced in treatment-naïve PBD patients but was present, even if less severe, in a relevant proportion of previously treated PDB cases, suggesting that frequently PDB patients are not correctly advised about the utility and beneficial effects of calcium and vitamin D supplementation during BP treatment. Similar observations concerning vitamin D deficiency were also evident in the interventional cohort, even though in this case the prevalence of vitamin D deficiency was even more pronounced likely because all the enrolled patients were naïve to BP treatment and possibly they had not been previously advised to receive calcium and vitamin D supplementation. Importantly, results of our study clearly underline that low 25OHD levels are associated with a higher prevalence of common acute adverse events occurring after the infusion of N-BP such hypocalcemia or APR in PDB, and that a preventive supplementation with vitamin D (cholecalciferol 50 000 IU/weekly for 8 weeks before N-BP infusion) in those subjects with circulating 25OHD levels below 20 ng/mL is able to prevent both adverse events in most cases. For ethical and safety issues we did not include a placebo arm in the interventional study and this may represent a potential limitation, since the prevalence of APR in cholecalciferol supplemented patients was compared with that observed in the overall cohort and in naïve cases from the observational cohort. However, results of our observational analysis were consistent with most of the evidence from the literature (41, 42), showing prevalence rates of APR in N-BP treated patients at first infusion around 40% to 50%, while in cholecalciferol treated patients from our study the occurrence of APR was remarkably reduced (below 10%). A similar reduced prevalence of APR was also evident in those patients from the observational study that received vitamin D supplementation and that had 25OHD levels above 20 ng/mL at the time of N-BP infusion.

Overall, these data confirm and extend evidence from previous studies in smaller cohorts of postmenopausal women or children undergoing intravenous infusion with N-BPs, where a higher prevalence of APR was observed in patients with vitamin D deficiency (31, 43, 44). In contrast, a retrospective analysis of 2 small trials with zoledronate in postmenopausal women with osteoporosis did not evidence any relationship between vitamin D status and the risk of APR (45). However, in that study different, nonconventional zoledronate dosages were used (with up to 60% of cases treated with low dosages of 1 mg or 2.5 mg) and most of the women were vitamin D replete, with 25OHD levels higher than 20 ng/mL in 94% of cases. This suggests that APR following N-BP treatment may indeed be related to vitamin D status, possibly through direct effects on γδ T cells, the subpopulations of T cells mainly involved in APR (31). Since we did not assess the T cells subpopulations in our cohorts, additional studies of circulating cell phenotypes should be required to further clarify this issue. However, an additive contribution of vitamin D replacement on the reduction of bone turnover and its potential implications on the occurrence of APR cannot be completely ruled out. Likewise, an adequate vitamin D status might contribute to obtain a less inflammatory bone phenotype for the bisphosphonates to act upon.

Moreover, it is also likely that the relationships between 25OHD levels and the occurrence of APR is not linear, being more evident when 25OHD levels are lower than 20 ng/mL. Indeed, in our cohort of patients APR was very common in the presence of severe vitamin D deficiency (25OHD < 10 ng/mL) affecting up to 80% of cases, even when patients already treated with N-BPs were considered. Thus, preventive vitamin D supplementation should be particularly indicated in patients with severe vitamin D deficiency in order to decrease the risk of APR. This might have important implications not only for the management of PDB, since APR was indicated as the main reason for discontinuing zoledronic acid treatment after a single infusion in more than 80% of osteoporotic patients (16).

Of interest, due to the large sample of cases included in the observational cohort, our data also allowed a better characterization of the occurrence of APR and its determinants in patients with PDB treated with different N-BP regimens. In keeping with most of the available literature, APR mainly occurred in those patients naïve from previous N-BP treatment, while was less prevalent among PDB cases that had been already treated with N-BPs. In the latter group of patients APR was less common in those cases treated with more potent N-BPs, while occurred at a same prevalence rate than in naïve patients in those cases treated with BPs without a nitrogen group, such as clodronate. Importantly, we also demonstrated that among patients already treated for PDB, APR occurs more frequently in those cases that reported a previous APR after N-BP treatment, suggesting that predisposing factors might also be involved in the pathogenesis of this complication. In this respect, the presence of SQSTM1 mutation was associated with higher prevalence rates of APR, including patients with a previous N-BP treatment for PDB, despite similar 25OHD levels. Indeed, SQSTM1 mutation carriers generally show a more severe PDB phenotype (46, 47).

In the overall group of patients, APR was also more common when N-BP treatment was given due to biochemical relapse, rather than when patients were treated because of PDB symptoms (mostly pain) in the presence of normal ALP levels. This might suggest an implication of bone turnover status before infusion on the occurrence of APR, at least in PDB.

In keeping with a previous report in a smaller sample (30), in both the observational and the interventional study we observed a rapid increase in PTH levels after N-BP infusion, which was more relevant in those patients experiencing an APR. Such an increase was associated with a parallel decrease in 25OHD, likely due to an enhanced 1α-hydroxylase activity, as well as with a transient decrease in Ca and P levels. This behavior was evidenced also in the interventional study, particularly in the 6 cases experiencing APR, despite vitamin D supplementation and the presence of 25OHD levels above the 30 ng/mL threshold in 5 out of 6 cases, suggesting that a global dysregulation of the vitamin D-PTH axis might be relevant for the occurrence of APR (eg, due to a long lasting vitamin D deficiency status). Indeed, a strong correlation between the rise in PTH levels and the inflammatory response of APR, as reflected by serum IL-6 levels, was also evident in the interventional study. This is in keeping with a previous report from Schweitzer et al. in a smaller sample of PDB cases (48). Remarkably, in the same study, an in vitro analysis on fetal mouse bone explants demonstrated that N-BPs such as olpandronate are able to suppress bone resorption but do not enhance IL-6 levels when given alone, while cotreatment with PTH resulted in a 6-fold stimulation of IL-6 release (48).

Finally, patients experiencing APR also showed a less relevant suppression of bone turnover markers after N-BP treatment than patients without APR, during the observation period.

Conclusions

In conclusion, our data better defined the characteristics and the risk factors of APR following N-BP treatment in PDB and disclosed for the first time that vitamin D deficiency is a relevant issue also in the management of PDB, affecting either treatment-naïve patients or cases already treated with N-BPs. Importantly, vitamin D supplementation and the achievement of adequate 25OHD levels should be recommended before intravenous infusion of N-BPs in order to minimize the risk of APR or hypocalcemia and possibly to maximize treatment response in terms of biochemical suppression of bone turnover at 12 months from infusion.

Abbreviations

Abbreviations
 
• 25OHD

  25-hydroxyvitamin D

 
• APR

  acute phase reaction

 
• BMI

  body mass index

 
• BP

  bisphosphonate

 
• CI

  confidence interval

 
• FPP

  farnesyl pyrophosphate

 
• IFN

  interferon

 
• IL

  interleukin

 
• IPP

  isoprenyl pyrophosphate

 
• IV

  intravenously

 
• PDB

  Paget’s disease of bone

 
• RR

  relative risk

 
• TNF

  tumor necrosis factor.

Acknowledgments

Financial Support: This work was supported by a Young Investigator research grant by the Italian Ministry of Health (GR-2011-02352160 to D. Merlotti).

Author Contributions: D.M. and L.G. Study conduct: D.M., D.R., T.P., M.A., R.M. Data collection: D.M., D.R., R.M., S.B., M.M., M.B.F., B.L. Data analysis: D.M., D.R., G.D., L.G. Data interpretation: D.M., D.R., G.D., L.G. Drafting manuscript: D.M. and L.G. Revising manuscript content: D.M., D.R., S.C., P.S., R.N., L.G. D.M. takes responsibility for the integrity of the data analysis. All authors read and approved the submitted version of the manuscript.

Additional Information

Disclosure Summary: D.R., R.M., T.P., M.A., M.M., S.B., M.B.F., B.L., S.C., and P.S. have nothing to declare. D.M. received honoraria from UCB Pharma. L.G. received honoraria from Sandoz. R.N. received honoraria from Sandoz and Pharma Group.

Data Availability: The datasets generated during and/or analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request.