chlorin-p6 and Mouth-Neoplasms

We investigated the anticancer chemotoxicity of previously reported iodinated chlorin p

Topics: Antineoplastic Agents; Apoptosis; Cell Line; Cell Line, Tumor; Cell Proliferation; Cell Survival; Coordination Complexes; Copper; Humans; Mouth Neoplasms; Porphyrins; Reactive Oxygen Species

Over-expression of histamine receptors has been reported in several types of malignancies. Earlier we have successfully demonstrated use of chlorin p6-histamine conjugate (Cp6-his) for improving cellular uptake and photo toxicity of Cp6 in oral cancer cell lines. In the present study, after having confirmed that histamine receptors are over-expressed in tumors of hamster cheek pouch, we investigated the efficacy of Cp6-his for photodynamic treatment (PDT) of tumors in this animal model.. Cp6-his (3mg/kg body weight) was injected intraperitoneally and its accumulation in tumor, surrounding tissue, normal mucosa and abdominal skin was monitored non-invasively by fluorescence spectroscopy. For PDT, tumors at 4h after Cp6-his administration were exposed to red light (660±25nm, 100J/cm(2)). Tumor damage and regression were assessed by histology and tumor volume measurements, respectively. Expression of histamine H2 receptors in tumor and normal mucosa was assessed by immuno-staining.. The accumulation of Cp6-his was higher in tumors as compared to normal mucosa at 4h after its administration. For Cp6 similar preferential accumulation was observed except that in normal mucosa the accumulation of Cp6 was more as compared to Cp6-his. The clearance of Cp6-his from skin was rapid showing ∼80% decrease within 48h from its peak level at 4h after drug injection. PDT led to extensive cellular damage and tumors of size up to ∼1000mm(3) regressed completely one week after PDT.. Higher tumor selectivity of Cp6-his and complete regression of bigger tumors after PDT suggest that conjugating Cp6 to histamine is a promising approach to improve PDT efficacy.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cheek; Cricetinae; Disease Models, Animal; Histamine; Humans; Male; Mesocricetus; Mouth Neoplasms; Photosensitizing Agents; Porphyrins; Treatment Outcome

Our previous studies in hamster cheek pouch model have shown that chlorin p (6) (Cp (6)), a chlorophyll derivative is a suitable photosensitizer for photodynamic treatment (PDT) of small tumors (<5 mm). However, for bigger tumors, the accumulation of Cp (6) was inadequate, which compromised the effectiveness of PDT. The purpose of present study was to investigate the possibility of improving the cellular uptake of Cp (6) by conjugating it to histamine, a biogenic amine that is known to modulate tumor growth and development via cell surface receptors.. The conjugate of Cp (6) and histamine (Cp (6)-his) was prepared by carbodiimide coupling reaction. Cellular uptake, intracellular localization and cytotoxicity of both Cp (6) and its conjugate were investigated in two human oral cancer cell lines (4451 and NT8e). The percentage of necrotic and apoptotic cells after PDT were also estimated using Hoechst 33342-propidium iodide staining.. In both the cell line, the cellular uptake of Cp (6)-his was found to be ~10 times higher when compared to Cp (6). Histamine led to a slight increase in intracellular uptake of Cp (6)-his, whereas ranitidine, a histamine H2 receptor antagonist, and incubation at lower temperature (~15°C) led to its inhibition, suggesting that uptake of Cp (6)-his is receptor mediated. Results on western blot confirmed the presence of H2 receptor in both the cell line. Observations on intracellular localization revealed that unlike Cp (6), which localized on multiple sites, Cp (6)-his showed localization on the cell membrane and around the perinuclear region. Moreover, the phototoxicity induced by Cp (6)-his was ~4 times higher when compared to Cp (6) in both the cell lines. There was, however, no significant difference in the mode of cell death.. Results suggest that conjugating Cp (6) with histamine can help improve the effectiveness of PDT in oral cancer cells by enhancing its intracellular delivery.

Topics: Apoptosis; Biological Transport; Carbodiimides; Cell Line, Tumor; Cell Membrane; Cell Nucleus; Cold Temperature; Cross-Linking Reagents; Histamine; Histamine H2 Antagonists; Humans; Kinetics; Mouth Neoplasms; Necrosis; Osmolar Concentration; Photochemotherapy; Photosensitizing Agents; Porphyrins; Ranitidine; Receptors, Histamine H2

We investigated tumor regression and the mode of tumor cell death induced by photodynamic treatment (PDT) with chlorin p(6) (Cp(6)) in hamster cheek pouch model of oral squamous cell carcinoma. Cp(6) was administered systemically through intraperitoneal injection and after 4h the tumors were subjected to photodynamic treatment using red light (660±25nm, fluence ∼100J/cm(2)). Tumor response to PDT was monitored by measuring the tumor volume before PDT and 1week after. Results show that smaller tumors (⩽80mm(3)) regressed completely after PDT with Cp(6) dose of 2.0mg/kg body weight and for the bigger tumors (∼180mm(3)) higher dose of Cp(6) (4.0mg/kg) was more effective. Tumors treated with lower Cp(6) dose showed infiltration of immune cells, absence of TUNEL labeling, smeared pattern of DNA fragmentation and no significant increase in caspase-3 activity suggestive of necrotic cell death and inflammation. In tumors treated with higher Cp(6) dose, features characteristic of apoptotic cell death such as extensive TUNEL positive labeling, increase in caspase-3 activity and laddered pattern of DNA fragmentation were observed and there was no infiltration of immune cells. PDT with Cp(6) was also found to lead to expression of matrix metalloprotease-9 (MMP-9) which was greater at lower drug dose PDT as compared to higher drug dose PDT. These results suggest that drug dose plays an important role in determining the mechanism of tumor cell death and effectiveness of PDT.

Topics: Animals; Carcinoma, Squamous Cell; Caspase 3; Cell Death; Cheek; Cricetinae; Dose-Response Relationship, Drug; Female; Models, Animal; Mouth Mucosa; Mouth Neoplasms; Photochemotherapy; Photosensitizing Agents; Porphyrins; Remission Induction; Tumor Burden

We studied pharmacokinetics and tumor response to photodynamic therapy (PDT) using chlorin p6 (CP6) in hamster cheek pouch model. CP6 was administered either intraperitoneally (IP) at a dose of 1.5 mg/kg body weight or applied topically at 1.0 mg/kg body weight and its accumulation in tumor, normal mucosa, and abdominal skin was measured by optical fiber-based fluorescence spectroscopy. Photodynamic therapy was performed by superficial illumination of tumor with 660 nm (+/-25 nm) light at a fluence rate of 100J/cm2 and tumor response to PDT was analyzed by histological examination. CP6 accumulation was higher in tumors as compared to adjoining tissue and normal mucosa at 4-6h after its IP administration. For relatively large tumors (size >8mm) topical application was observed to be more effective than IP. The level of CP6 in tumor, surrounding tissue, normal mucosa and skin was seen to decrease rapidly within 24h after its administration and was undetectable at longer time (>72 h) intervals. PDT of small tumors at 4h after IP injection of CP6 resulted in complete tumor necrosis. Whereas, PDT of large tumors receiving CP6 topically caused necrosis in 300-800 microm superficial region of the tumor. In one animal kept for follow up in each treatment group, it was observed that small tumors disappeared completely leaving scar tissue, while large tumor had significant reduction in tumor size. The use of CP6 for PDT of oral cancer is suggested.

Topics: Animals; Carcinoma, Squamous Cell; Cheek; Cricetinae; Drug Screening Assays, Antitumor; Male; Models, Animal; Mouth Mucosa; Mouth Neoplasms; Photochemotherapy; Photosensitizing Agents; Porphyrins