A significant literature exists on NRTIs that have implicated mitochondrial dysfunction, reviewed in.44
The inhibition of mitochondrial polymerase gamma could occur by competitive inhibition of polymerase gamma and/or the incorporation and subsequent DNA chain termination. The result would be a reduction in mitochondrial DNA synthesis. NRTIs could also indirectly inhibit DNA synthesis by inducing nucleotide pool imbalances, oxidative stress, and DNA damage. Both direct and indirect mechanisms would result in the alteration of mitochondrial gene expression, which would cause mitochondrial dysfunction. NRTIs have greater affinity for affecting different tissue types as well as the nature of the mitochondrial-induced dysfunction in these tissue types, depending upon the NRTI. The analysis of NRTI-associated cellular stress has indicated that the mechanisms involved can be quite complex. NRTIs can result in altered gene expression profiles.50
Altered gene expression profiles have been observed in the absence of mitochondrial DNA depletion, suggesting that NRTIs can cause mitochondrial dysfunction and not inhibit mitochondrial DNA polymerase gamma.52
The association of tenofovir with mitochondrial dysfunction has been investigated. In general, no mitochondrial dysfunction was observed with tenofovir.53
Other studies have reported a lowering of mitochondrial dysfunction when drug regimens were changed and NRTIs were replaced with tenofovir.57
A recent study in rats found that very high doses (ie, 100 mg/kg/day) of tenofovir reduced mitochondrial DNA and gene expression in kidney cells.60
The relevance of this observation given the very high dosing used is unclear. In stark contrast, in an HIV transgenic mouse model tenofovir treatment was found to increase mitochondrial DNA content in kidney cells.61
Further analysis of mitochondrial DNA obtained from renal proximal tubules microdissected from kidney sections suggested that tenofovir lowered mitochondrial DNA levels in the renal proximal tubules.61
To date, no studies of mitochondrial dysfunction being associated with tenofovir-mediated bone loss have been reported. Overall, the current literature does not provide strong evidence for mitochondrial dysfunction being a major mechanism for tenfovir-mediated cellular stress, particularly in regards to bone loss.
In vitro studies have previously reported that tenofovir diphosphate is a poor substrate and weak inhibitor of rat DNA polymerase alpha, delta and epsilon.55
Combined with the report of tenofovir diphosphate being poorly incorporated into DNA by the human mitochondrial DNA polymerase gamma,49
data are limited that would support a direct role of tenofovir in bone loss by the inhibition of either nuclear or mitochondrial DNA replication. While data are lacking for tenofovir diphosphate being a good substrate for cellular DNA polymerases, the in vivo data correlating tenofovir therapy with bone loss, particularly in children and adolescents who have very active and ongoing bone growth, implicate a role for tenofovir’s ability to impact cellular DNA synthesis. Such an impact could be either direct or indirect, such as 1) incorporation and DNA chain termination, 2) DNA damage, 3) alteration of deoxynucleotide transport, and/or 4) nucleotide pool imbalances, which would perturb cellular DNA synthesis. The perturbation of cellular (ie, nuclear and/or mitochondrial) DNA synthesis would result in altered gene expression. The alteration of gene expression for genes involved in bone maintenance could explain the clinical observation of bone density loss during tenofovir treatment.
We propose three potential mechanisms for tenofovir-associated bone loss. These include 1) preferential uptake by osteoclasts (altering gene expression and resulting in increased bone resorption), 2) update by osteoblasts (altering gene expression and decreasing bone formation), and 3) uptake by both osteoclasts and osteoblasts (altering gene expression of both cells types and ultimately the balance between bone resorption and bone formation – resulting in bone loss).
Since TDF is a phosphonate, it is possible that this could enhance its uptake into cells (), which would increase the probability of cellular stress. Bisphosphonates (ie, diphosphonates) are drugs used clinically to prevent loss of bone density, particularly in diseases such as osteoporosis, bone metastasis, multiple myeloma, Paget’s disease of bone, and primary hyperparathyroidism. Bisphosphonates target bone and inhibit osteoclast function after their cellular uptake by inducing apoptosis.62
Since tenofovir and TDF are both phosphonates, it is conceivable that they could also have an association with bone and be selectively taken up by osteoclasts by a mechanism similar to that of bisphosphonates, ultimately causing cellular stress. The resulting cellular stress would likely perturb cellular DNA synthesis (ie, nuclear and/or mitochondrial) and gene expression (). For example, the reduction of gene expression for an osteoclast gene that is involved in signaling osteoblast activity could ultimately result in a loss of bone density. It is formally possible that adjuvant treatment of bone density loss could improve the durability of tenofovir-containing HAART. In the case of osteoclast hyperactivity, it is possible that bisphosphonates could reduce bone density loss associated with tenofovir treatment.
Figure 2 The osteoclast as a target for TDF. A) Bone tissue, osteoblasts and osteoclasts. TDF, as a phosphonate, associates with bone tissue. Bone resorption by osteoclasts would result in the preferential uptake of TDF. B) Impact of TDF on osteoclast DNA synthesis (more ...)
The loss of bone density due to TDF exposure could also be associated with tenofovir-induced renal dysfunction – particularly renal proximal tubule dysfunction.42
The failure of renal proximal tubular cells to reabsorb filtered bicarbonate from the urine would result in urinary bicarbonate wasting and subsequent acidemia and a more general dysfunction of the proximal tubular cells – called Fanconi’s syndrome. Commonly observed conditions in Fanconi syndrome include aminoaciduria, glycosuria, tubular proteinuria, and uricosuria. Importantly, the main clinical feature of Fanconi’s syndrome is bone demineralization (osteomalacia or rickets) due to phosphate wasting. Therefore, TDF-associated bone density loss may an outcome of renal dysfunction.