Zoledronic acid induces formation of a pro-apoptotic ATP analogue and isopentenyl pyrophosphate in osteoclasts in vivo and in MCF-7 cells in vitro

Background and purpose:

Bisphosphonates (BPs) are highly effective inhibitors of bone resorption. Nitrogen-containing bisphosphonates (N-BPs), such as zoledronic acid, induce the formation of a novel ATP analogue (1-adenosin-5′-yl ester 3-(3-methylbut-3-enyl) ester triphosphoric acid; ApppI), as a consequence of the inhibition of farnesyl pyrophosphate synthase and the accumulation of isopentenyl pyrophosphate (IPP). ApppI induces apoptosis, as do comparable metabolites of non-nitrogen-containing bisphosphonates (non-N-BPs). In order to further evaluate a pharmacological role for ApppI, we obtained more detailed data on IPP/ApppI formation in vivo and in vitro. Additionally, zoledronic acid-induced ApppI formation from IPP was compared with the metabolism of clodronate (a non-N-BP) to adenosine 5′(β,γ-dichloromethylene) triphosphate (AppCCl₂p).

Experimental approach:

After giving zoledronic acid in vivo to rabbits, IPP/ApppI formation and accumulation was assessed in isolated osteoclasts. The formation of ApppI from IPP was compared with the metabolism of clodronate in MCF-7 cells in vitro. IPP/ApppI and AppCCl₂p levels in cell extracts were analysed by mass spectrometry.

Key results:

Isopentenyl pyrophosphate/ApppI were formed in osteoclasts in vivo, after a single, clinically relevant dose of zoledronic acid. Furthermore, exposure of MCF-7 cells in vitro to zoledronic acid at varying times and concentrations induced time- and dose-dependent accumulation of IPP/ApppI. One hour pulse treatment was sufficient to cause IPP accumulation and subsequent ApppI formation, or the metabolism of clodronate into AppCCl₂p.

Conclusions and implications:

This study provided the first conclusive evidence that pro-apoptotic ApppI is a biologically significant molecule, and demonstrated that IPP/ApppI analysis is a sensitive tool for investigating pathways involved in BP action.     

Safety evaluation of an IPP tripeptide-containing milk protein hydrolysate

Tensguard™ is a milk protein hydrolysate containing the lactotripeptide IPP. It is derived from cow’s milk, which is present in the human diet and has a safe history of consumption. The final Tensguard™ product, a supplement or a functional food ingredient, is intended for use by people who want to follow a healthy diet and lifestyle in order to manage their blood pressure. The safety-in-use of commercial lactotripeptide-containing products has been confirmed in several in vitro and in vivo toxicity studies and in studies with humans. To support the safety, Tensguard™ was examined in three in vitro genotoxicity tests (bacterial reverse mutation test, mammalian cell gene mutation test and mammalian chromosomal aberration test) and in a 90-day repeated-dose oral toxicity study in rats. The genotoxicity tests confirm that Tensguard™ is not mutagenic or clastogenic. The NOAEL from the 90-day study was at the highest dose tested, i.e. 4% in the diet. The NOAEL is equivalent to an overall mean intake of 2 g Tensguard™/kg body weight/day and corresponds to 40 mg IPP/kg body weight/day. This is 141-fold higher than the maximal anticipated intake. In conclusion, Tensguard™ is safe under the conditions of intended use.     

The effects of systemic treatment with aminobisphosphonates and statins on circulating Vγ9Vδ2-T cells in patients with advanced cancer

Aminobisphosphonates (NBP) are used for treatment of metastatic bone disease. Frequently, patients undergoing NBP-treatment experience side-effects, known as acute phase response (APR), resulting from cytokine production by Vγ9Vδ2-T cells. As opposed to NBP, statins reduce intracellular phosphoantigen levels and prevent NBP-induced Vγ9Vδ2-T cell activation in vitro. We conducted a pilot study in patients with (bone-)metastasized malignancies receiving NBP-treatment and evaluated the phenotype and function of circulating Vγ9Vδ2-T cells in vivo and the effects of statins on Vγ9Vδ2-T cell responses and the associated APR. We observed reduced expression of perforin, granzyme B and HLA-DR on Vγ9Vδ2-T cells in patients treated with NBP and statins. However, statins could not prevent NBP-induced changes in circulating Vγ9Vδ2-T cell numbers or production of IFNγ and TNFα. Consistent with this, simvastatin could not prevent the occurrence of APR upon NBP-infusion. These observations call for the exploration of alternative strategies to prevent collateral APR upon NBP treatment.     

PINCH‐2 presents functional copy number variation and suppresses migration of colon cancer cells by paracrine activity

In recent years, characterization of cancer and its environment has become necessary. However, studies of the cancer microenvironment remain insufficient. Copy number variations (CNVs) occur in 40% of cancer‐related genes, but few studies have reported the correlation between CNVs in morphologically normal tissues adjacent to cancer and cancer progression. In this study, we evaluated cancer cell migration and invasion according to the genetic differences between cancer tissues and their surrounding normal tissues. To study the field cancerization effect, we screened 89 systemic metastasis‐related CNVs from morphologically normal tissues adjacent to colon cancers. Among these CNVs, LIM and senescent cell antigen‐like domain 2 (PINCH‐2) showed copy number amplification and upregulation of mRNA in the nonrelapsed group compared to the systemic relapse group. PINCH‐2 expression in colon cancer cells was lower than that in normal epithelial colon cells at both the protein and mRNA levels. Suppression of PINCH‐2 resulted in decreased formation of the PINCH‐2‐IPP (PINCH‐2, integrin‐linked kinase and α‐parvin) complex and reciprocally increased formation of the PINCH‐1‐IPP complex. Although PINCH‐2 expression of survival pathway‐related proteins (Akt and phospho‐Akt) did not change upon suppression of PINCH‐2 expression, cell migration‐related proteins [matrix‐metalloproteinase (MMP)?9 and ?11] were upregulated through autocrine and paracrine activation. Thus, PINCH‐2 participates in decreased systemic recurrence by competitively regulating IPP complex formation with PINCH‐1, thereby suppressing autocrine and paracrine effects on motility in colon cancer. This genetic change in morphologically normal tissue suggests a field cancerization effect of the tumor microenvironment in cancer progression.     

Penile implant infection factors: a contemporary narrative review of literature

ObjectiveWe aim to review and summarize published literature that features implanted penile devices and details infection of these devices as a complication. In particular, we will detail the factors that influence infection of penile implants.BackgroundTypes of penile prostheses (PP) include inflatable implants and semirigid implants; these are utilized for treatment of erectile dysfunction. Likely the most feared complication of penile implants is infection. There are a handful of factors that are implicated in device infection.MethodsSearches were performed using MEDLINE and PubMed databases using keywords and phrases ‘penile implant AND infection’; ‘penile prosthesis AND infection’; ‘penile implant infection’. We have presented results from our literature search. We divided these into ‘Surgical Elements’ and ‘Patient Selection and Factors.’ Each topic is discussed in its own section.ConclusionsStrides have been made since the initial penile prosthesis (IPP) surgeries to improve infection rates including diabetes control, antibiotic coating of devices, and antibiotic implementation. Going forward, more studies, especially randomized control trials, need to focus on defining levels of diabetic control (sugar control and A1C control), determining the role of metabolic syndrome in infection promotion and determining laboratory values which could be predictive of infection. We present a discussion of important factors to consider in the realm of PP infections. In addition, we include studies which discuss topics for future directions in decreasing the number of infections seen with PP.     

Sensitization of Cervical Carcinoma Cells to Paclitaxel by an IPP5 Active Mutant

Paclitaxel is one of the best anticancer agents that has been isolated from plants, but its major disadvantageis its dose-limiting toxicity. In this study, we obtained evidence that the active mutant IPP5 (8-60hIPP5m), thelatest member of the inhibitory molecules for protein phosphatase 1, sensitizes human cervix carcinoma cellsHeLa more efficiently to the therapeutic effects of paclitaxel. The combination of 8-60hIPP5m with paclitaxelaugmented anticancer effects as compared to paclitaxel alone as evidenced by reduced DNA synthesis andincreased cytotoxicity in HeLa cells. Furthermore, our results revealed that 8-60hIPP5m enhances paclitaxelinducedG2/M arrest and apoptosis, and augments paclitaxel-induced activation of caspases and release ofcytochrome C. Evaluation of signaling pathways indicated that this synergism was in part related to downregulationof NF-ҝB activation and serine/threonine kinase Akt pathways. We noted that 8-60hIPP5m downregulatedthe paclitaxel-induced NF-ҝB activation, IқBα degradation, PI3-K activity and phosphorylation ofthe serine/threonine kinase Akt, a survival signal which in many instances is regulated by NF-ҝB. Together, ourobservations indicate that paclitaxel in combination with 8-60hIPP5m may provide a therapeutic advantage forthe treatment of human cervical carcinoma