Adjuvant Photodynamic Therapy, Mediated via Topical Versus Systemic Administration of 5-Aminolevulinic Acid for Control of Murine Mammary Tumor after Surgical Resection.

Treatment de-escalation is sought in the management of precursor lesions of early stage breast cancer, driving the appeal of adjuvant modalities to lumpectomy that reduce toxicity and minimally detract from patient quality of life. We investigate photodynamic therapy (PDT), with the photosensitizing prodrug, 5-aminolevulinic acid (ALA), as adjuvant therapy to complete resection of murine mammary tumor (propagated from TUBO cells). ALA was delivered either systemically (oral, 250 mg kg-1 ) at 5 h before 632 nm illumination or topically (20% solution) to the resection site at 10 min before light delivery to 135 J cm-2 . Treatment with either oral-ALA-PDT (oALA-PDT) or topical-ALA-PDT (tALA-PDT) to the mammary fat pad after TUBO complete resection (CR) produced long-term tumor control with 90-day complete response rates of 21% and 32%, respectively, compared to control rates of 0-5% in mice receiving only CR. Thus, CR/tALA-PDT was equipotent to CR/oALA-PDT despite ~10-fold lower levels of ALA-induced protoporphyrin XI as photosensitizer after topical versus oral-ALA administration. CR/oALA-PDT produced more vascular damage, greater proportion of tissue-resident neutrophils and stronger inflammation when compared to CR/tALA-PDT. Collectively, these data provide rationale for ongoing investigation of ALA-PDT as adjuvant therapy after lumpectomy for increased probability of local control in the treatment of breast cancer.

Light Fluence Rate and Tissue Oxygenation (St O2 ) Distributions Within the Thoracic Cavity of Patients Receiving Intraoperative Photodynamic Therapy for Malignant Pleural Mesothelioma.

The distributions of light and tissue oxygenation (St O2 ) within the chest cavity were determined for 15 subjects undergoing macroscopic complete resection followed by intraoperative photodynamic therapy (PDT) as part of a clinical trial for the treatment of malignant pleural mesothelioma (MPM). Over the course of light delivery, detectors at each of eight different sites recorded exposure to variable fluence rate. Nevertheless, the treatment-averaged fluence rate was similar among sites, ranging from a median of 40-61 mW cm-2 during periods of light exposure to a detector. St O2 at each tissue site varied by subject, but posterior mediastinum and posterior sulcus were the most consistently well oxygenated (median St O2 >90%; interquartile ranges ~85-95%). PDT effect on St O2 was characterized as the St O2 ratio (post-PDT St O2 /pre-PDT St O2 ). High St O2 pre-PDT was significantly associated with oxygen depletion (St O2 ratio < 1), although the extent of oxygen depletion was mild (median St O2 ratio of 0.8). Overall, PDT of the thoracic cavity resulted in moderate treatment-averaged fluence rate that was consistent among treated tissue sites, despite instantaneous exposure to high fluence rate. Mild oxygen depletion after PDT was experienced at tissue sites with high pre-PDT St O2 , which may suggest the presence of a treatment effect.

Luminol Chemiluminescence Reports Photodynamic Therapy-Generated Neutrophil Activity In Vivo and Serves as a Biomarker of Therapeutic Efficacy.

Inflammatory cells, most especially neutrophils, can be a necessary component of the antitumor activity occurring after administration of photodynamic therapy. Generation of neutrophil responses has been suggested to be particularly important in instances when the delivered photodynamic therapy (PDT) dose is insufficient. In these cases, the release of neutrophil granules and engagement of antitumor immunity may play an important role in eliminating residual disease. Herein, we utilize in vivo imaging of luminol chemiluminescence to noninvasively monitor neutrophil activation after PDT administration. Studies were performed in the AB12 murine model of mesothelioma, treated with Photofrin-PDT. Luminol-generated chemiluminescence increased transiently 1 h after PDT, followed by a subsequent decrease at 4 h after PDT. The production of luminol signal was not associated with the influx of Ly6G+ cells, but was related to oxidative burst, as an indicator of neutrophil function. Most importantly, greater levels of luminol chemiluminescence 1 h after PDT were prognostic of a complete response at 90 days after PDT. Taken together, this research supports an important role for early activity by Ly6G+ cells in the generation of long-term PDT responses in mesothelioma, and it points to luminol chemiluminescence as a potentially useful approach for preclinical monitoring of neutrophil activation by PDT.

Fluence Rate Differences in Photodynamic Therapy Efficacy and Activation of Epidermal Growth Factor Receptor after Treatment of the Tumor-Involved Murine Thoracic Cavity.

Photodynamic therapy (PDT) of the thoracic cavity can be performed in conjunction with surgery to treat cancers of the lung and its pleura. However, illumination of the cavity results in tissue exposure to a broad range of fluence rates. In a murine model of intrathoracic PDT, we studied the efficacy of 2-(1-hexyloxyethyl)-2-devinyl pyropheophorbide-a (HPPH; Photochlor(®))-mediated PDT in reducing the burden of non-small cell lung cancer for treatments performed at different incident fluence rates (75 versus 150 mW/cm). To better understand a role for growth factor signaling in disease progression after intrathoracic PDT, the expression and activation of epidermal growth factor receptor (EGFR) was evaluated in areas of post-treatment proliferation. The low fluence rate of 75 mW/cm produced the largest reductions in tumor burden. Bioluminescent imaging and histological staining for cell proliferation (anti-Ki-67) identified areas of disease progression at both fluence rates after PDT. However, increased EGFR activation in proliferative areas was detected only after treatment at the higher fluence rate of 150 mW/cm. These data suggest that fluence rate may affect the activation of survival factors, such as EGFR, and weaker activation at lower fluence rate could contribute to a smaller tumor burden after PDT at 75 mW/cm.

Tumor vascular microenvironment determines responsiveness to photodynamic therapy.

The efficacy of photodynamic therapy (PDT) depends upon the delivery of both photosensitizing drug and oxygen. In this study, we hypothesized that local vascular microenvironment is a determinant of tumor response to PDT. Tumor vascularization and its basement membrane (collagen) were studied as a function of supplementation with basement membrane matrix (Matrigel) at the time of tumor cell inoculation. Effects on vascular composition with consequences to tumor hypoxia, photosensitizer uptake, and PDT response were measured. Matrigel-supplemented tumors developed more normalized vasculature, composed of smaller and more uniformly spaced blood vessels than their unsupplemented counterparts, but these changes did not affect tumor oxygenation or PDT-mediated direct cytotoxicity. However, PDT-induced vascular damage increased in Matrigel-supplemented tumors, following an affinity of the photosensitizer Photofrin for collagen-containing vascular basement membrane coupled with increased collagen content in these tumors. The more highly collagenated tumors showed more vascular congestion and ischemia after PDT, along with a higher probability of curative outcome that was collagen dependent. In the presence of photosensitizer-collagen localization, PDT effects on collagen were evidenced by a decrease in its association with vessels. Together, our findings show that photosensitizer localization to collagen increases vascular damage and improves treatment efficacy in tumors with greater collagen content. The vascular basement membrane is thus identified to be a determinant of therapeutic outcome in PDT of tumors.