Vitamin D₃ cryosensitization increases prostate cancer susceptibility to cryoablation via mitochondrial‐mediated apoptosis and necrosis

Abstract

Objectives To investigate the effect and molecular mechanisms of action of Vitamin D3 (VD3) as a neo-adjunctive agent before cryosurgery in an effort to increase treatment efficacy for prostate cancer (CaP). To eliminate the potential for disease recurrence that exists at the periphery of the freeze lesion, where temperatures may be insufficient to destroy both androgen-sensitive (AS) and androgen-insensitive (AI) CaP. Methods Human CaP cells, LNCaP, were each genetically altered to express the AS and AI phenotypes and subjected to VD3 treatment and freezing in an in vitro and tissue-engineered model. Cell viability, caspase inhibitor and western blot studies were used to determine the basis of the different responses of AI and AS cells to VD3 cryosensitization. Results VD3 was found to be a highly effective cryosensitizer, resulting in a >50% overall increase in cell death after -15°C freezing. Fluorescence microscopy, western blot analysis and caspase protease assays confirmed that the increased activation of apoptosis was modulated through a mitochondrial-mediated pathway. Caspase inhibition studies showed that apoptosis played an integral role in cell death, with VD3 cryosensitivation-induced apoptotic events responsible for > 30% of the overall cell death after -15°C freezing. Conclusions The present study suggests that the use of VD3 as a cryosensitizer increases cryoablation efficacy through the increased activity of apoptosis as well as through necrosis. The data show that through VD3 treatment the overall level of AI CaP cell tolerance to freezing is reduced to a level similar to that of AS CaP. VD3 pre-treatment in conjunction with cryoablation may increase treatment efficacy and reduce disease recurrence for CaP patients.

Evaluation of a new epicardial cryoablation system for the treatment of Cardiac Tachyarrhythmias

“…It is important to understand the model utilized and recognize the benefits and limitations in order to frame the results. Several studies have shown that the distribution of the -20°C and -40°C critical isotherms is often restricted to the center of the frozen mass (closest to the cryoprobe), thereby comprising less than 35% and 20% of the frozen mass, respectively (53)(54)(55)58,61,67) The typical freeze times reported in these and other studies range from 5 to 10 minutes which are applied in multiple (repeat) freeze/ thaw scenarios. While limited, these data provide a reference point for comparison of modern technologies.…”

“…The data presented herein suggests that the SCN epicardial cryoprobe provides for the rapid delivery of an effective cryoablative dose to the target tissue using the ex vivo models evaluated. The data demonstrate that the delivery of a "thermal dose" of -30°C [59,[60][61][62][63][64][65][66][67][68] across tissue ranging from 5 mm to 1 cm in thickness under high (37°C) circulating heat loads in less than 2 minutes. Analysis of the thermal spread of the lethal temperatures during a 2-minute freeze was limited to within 5 mm of the probe surface thereby resulting in the creation of a defined linear ablation zone.…”

Mechanisms of cryoablation: Clinical consequences on malignant tumors

“…Antifreeze proteins have been shown to affect the rate and morphology of ice crystal formation and increase cell destruction in a prostate cancer grown in mice and rats [84,99]. Vitamin D 3 has been shown in cell culture and animal studies to be a highly effective cryosensitizing agent [12]. While diverse in nature and specific mechanisms (pathways) of action, a common theme of enhanced therapeutic outcome has emerged through the use of adjunctive pretreatment of cancer prior to cryoablation.…”

“…Through various in vitro , in vivo , and clinical reports, improved cancer destruction at mild sub-freezing temperatures following brief low dose exposure to general or specific apoptotic inducing agents has been shown. In some cases, such as the use of vitamin D 3 as a sensitizing agent, complete prostate cancer cell death has been demonstrated following freezing to temperatures as warm as −10°C in both in vitro and in vivo models [12,61,111]. …”

“…The diverse techniques have been broadly divided into thermal and non-thermal methods. The mechanisms of cryoablative damage are more completely described when compared to heat-based procedures [11,12,61,111]. …”