Aminolevulinate-based photodynamic therapy (ALA-PDT) is an effective treatment for cutaneous pre-cancers (Actinic Keratoses; AK) and Basal Cell Carcinoma (BCC), the most common skin malignancies. When administered in a conventional regimen with 1-4 h of ALA preincubation prior to light exposure, ALA-PDT elicits stinging pain during illumination that patients find objectionable. To avoid this pain, we have described a new regimen called metronomic PDT (mPDT) which is similar to daylight PDT but uses blue light (Kaw et al, J Am Acad Dermatol 2019). Metronomic PDT is not only painless but also nearly as effective as conventional PDT for AK lesion clearance. In this investigation, murine models of AK induced by repeated UVB exposure were treated with mPDT, followed by time-course analyses of immune responses in the lesions harvested. Our preliminary data showed that relative to conventional PDT, cell death (apoptosis) and generation of Reactive Oxygen Species (ROS) were compromised in mPDT samples. However, relative to untreated controls, enhanced recruitment/infiltration of immune cells that mediate innate immunity [neutrophils (Ly6G+) and macrophages (F4/80+)] was observed at early times after mPDT. Just as importantly, enhanced presence of cells regulating adaptive immune responses [T cells (CD3+, CD8+ and Foxp3+)] was observed at later times post mPDT. Activation of calreticulin and HMGB1 (markers of Damage Associated Molecular Patterns, DAMPs) were also observed in mPDT treated lesions. Our results suggest that mPDT can be just as effective as conventional PDT for treatment of skin cancer and pre-cancer, and that the therapeutic mechanisms may involve immune cell responses triggered by metronomic PDT.
Aminolevulinate-based photodynamic therapy (ALA-PDT) is effective for actinic keratosis (AK), but when administered in a conventional regimen, PDT can elicit pain during illumination that is very uncomfortable for patients. In conventional PDT, a 1-4 hr preincubation period occurs after ALA application, prior to the start of illumination. In a new regimen called metronomic PDT (mPDT), which simulates daylight PDT, blue light (405 nm) is delivered concurrently with ALA application and thereby provides a treatment that is not only painless but also nearly as effective as conventional PDT for AK lesion clearance. In this investigation, a murine AK model, generated by repeated UVB exposure, was treated with either mPDT or PDT. Lesion clearance was monitored via area measurements, and samples were harvested for assessment of biological mechanisms. Compared to pretreatment (100%), the average lesion area decreased to 47% and 32% in PDT, and to 57% and 40% in mPDT at 1 and 2 weeks after PDT, respectively. Relative to untreated controls, we observed enhanced cell death (by H&E staining, apoptosis, and TUNEL assay), generation of Reactive Oxygen Species (ROS, by CM-H2DCFDA staining), and autophagy (by Atg 5 and Atg 7 expression) in both the PDT and mPDT samples. Cleaved (activated) Caspase-3 was specifically observed only in PDT samples. Immunomodulation by inflammatory cells was indicated by enhanced infiltration/retention of neutrophils and macrophages in mPDT samples. Our results suggest that mPDT can be just as effective as conventional PDT for treatment of AK, but the mechanisms may be quite different.
Breast cancer (BCA) in women is a leading cause of mortality and morbidity with distant metastases occurring in ~40% of cases. Here, as an alternative to ionizing radiation therapy and chemotherapy and their associated side effects, we explored a new combination approach using capecitabine (CPBN) and aminolevulinate-based photodynamic therapy (PDT). We have recently shown that a combination PDT approach in which 5-fluorouracil (5FU), a differentiation-promoting agent, increases the levels of PpIX in cancer cells when given as a neoadjuvant prior to ALA. However, 5FU can be toxic when administered systemically at high levels. We reasoned that CPBN, a known chemotherapeutic drug for BCA and less toxic than 5FU (because CPBN is metabolized to 5FU specifically within tumor tissues), might work equally well as a PDT neoadjuvant. Murine 4T1 BCA cells harboring a luciferase transgene were injected into breast fat pads of female nude mice. CPBN (600 mg/kg/day) was administered by oral gavage for 3 days followed by intraperitoneal ALA administration and PDT with red light (633 nm) on day 4. Tumor growth and regression were monitored in vivo using bioluminescence imaging. Histological changes in primary tumors and metastases were assessed after necropsy. CPBN pretreatment of 4T1 tumors increased cellular differentiation, reduced proliferation, raised PpIX levels, enhanced tumor cell death, and reduced metastatic spread (incidence and tumor load) of 4T1 cells post-PDT, relative to vehicle-only controls. The use of CPBN as a non-toxic PDT neoadjuvant for treatment of BCA represents a novel approach with significant potential for translation into the clinic.
Aminolevulinic acid based photodynamic therapy (ALA-PDT) is a popular and efficacious treatment for actinic keratosis (AK). However, standard PDT can elicit stinging pain during illumination, and hence is not always favored by patients. In a new regimen called metronomic PDT (mPDT), similar to daylight PDT but using blue light, the illumination is delivered concurrently with ALA application rather than after a 1-hour pre-incubation (conventional regimen, PDT). In the clinic, mPDT is not only painless but also nearly as effective as PDT for AK lesion clearance. In this investigation, a murine AK model (generated by repeated UVB exposure) was treated with either mPDT or PDT. Lesion clearance was followed by area measurement, and samples were harvested for mechanistic analyses. Compared to pretreatment (100%), the average lesion area was reduced to 47% and 32% in PDT, and to 57% and 40% in mPDT at 1- and 2-weeks post PDT, respectively. Relative to untreated controls, enhanced cell death (histomorphology by H&E staining and apoptosis by TUNEL assay), and generation of Reactive Oxygen Species (ROS; CM-H2DCFDA staining) were observed in both PDT and mPDT samples. Activation of cleaved Caspase-3 was specifically observed only in PDT samples. Immunomodulation by inflammatory cells was observed by enhanced infiltration/retention of neutrophils and macrophages in metronomic PDT samples. Our results suggest that metronomic PDT can be just as effective as conventional PDT for treatment of AK, but the mechanisms may be quite different.
Breast cancer (BCA) is the most frequently diagnosed cancer in women, with distant metastases to lung, liver, bone and skin occurring in approximately 40% of cases. Radiation therapy (RT) has been successfully employed for the treatment of BCA; however, multiple rounds of RT are associated with undesirable cutaneous side effects. This study explores PDT as a therapeutic alternative, to be given alone or in combination with RT and chemotherapy. Earlier, we had developed differentiation-enhanced combination photodynamic therapy (cPDT) using a neoadjuvant (5-fluorouracil; 5FU) prior to PDT. The neoadjuvant increases the levels of PpIX, leading to better efficacy following aminolevulinate (ALA)- based PDT. Here, to avoid the toxicity of systemic 5FU, we used a nontoxic 5FU precursor (Capecitabine; CPBN) in a new cPDT regimen. CBPN, a standard chemotherapeutic for BCA, is metabolized to 5FU specifically within tumor tissue. Murine (4T1) BCA cells were injected into breast fat pads of nude mice. CPBN was administered by oral gavage followed by intraperitoneal ALA and red light for PDT. CPBN pretreatment of 4T1 tumors led to increased tumor cell differentiation (3.5 fold), homogenous elevation of intratumoral PpIX levels (4.5 fold), and enhanced tumor cell death post-PDT (5 fold), relative to vehicle control. Using an in vivo imaging system (IVIS), a decline in tumor growth following CPBN-PDT was observed. Results showing the effect of CPBN-PDT on distant metastases of BCA to lung, lymph nodes and skin will be presented. In summary, CPBN-PDT, a novel combination approach, has a significant potential for translation into the clinic.
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