Chlorin E6

Ce6 which is the shortened form of Chlorin E6, also known as Phytochlorin.

The molecular structure of Chlorin E6 is diagrammatically presented in Figure 1. Its molecular structure contains three carboxylic groups. The complete synthesis process for Chlorin E6 is described in the scientific literature in detail.

Figure 1 The Molecular Structure of Chlorin E6

Chemically, Chlorin E6 is a second-generation derivative of porphyrin which in turn a class of organic compounds. Being a porphyrin derivative, Chlorin E6 can acts as sensitizer and thus initiates the production of Reactive Oxygen Species ROS in the treated cells upon exposure to the stimuli, i.e., light. This high amount of the Reactive Oxygen Species ROS leads the treated cells to the cell cycle arrent, autophagy or apoptosis which is a type of programmed cell death via the respective signaling pathways. That is how Chlorin E6 is used for the treatment of tumor cells in a method named as Photodynamic therapy (PDT). The overall concept of this treatment procedure is schematically presented in Figure 2. This treatment is completely non-invasive.

Figure 2 The core concept of the novel non-invasive cancer therapy treatment, i.e., Photodynamic therapy (PDT). The targeted cells are exposed to light; and as result the targeted cells are either killed via autophagy/ apoptosis or prevented from proliferation by arresting them at specific stages of the cell cycle. Here in this figure, if the stimulus is ultrasound waves instead of light, the same procedure would be called as Sonodynamic therapy (SDT).

On the molecular level, the underlying mechanism that is driving the process of Photodynamic therapy (PDT) is schematically depicted in Figure 3 and Figure 4, where PS and PS* represent the normal state and excited state of the Chlorin E6, respectively. In Figure 4, porphyrin is used as a general term and it actually represents the Chlorin E6 in this case.  In this mechanism, as the targeted cells are exposed to the light; the molecules of the Photosensitive compound PS, which are in this case are of the Chlorin E6, absorb that light and are excited to PS*. The excited state, i.e., PS* is chemically unstable, therefore these PS* molecules return back to their original state, i.e., PS. As the PS* molecules return, not only fluorescence along with the heat is released but also Reactive Oxygen Species ROS is produced either through Type 1 photochemistry or Type 2 photochemistry. In the Type 1 photochemistry, PS* reacts with its legitimate ligand and leads to the production free radicals, followed by the production of the Reactive Oxygen Species ROS. In other case, in the Type 2 photochemistry, PS* reacts with the oxygen in the molecular form having triplet state ³O₂. This reaction involves the transfer of spin-changing energy. By this pathway, singlet oxygen is produced which a very strong oxidizing agent and have a key role in the anti-tumor activity. These produced Reactive Oxygen Species ROS leads to either the inhibited proliferation in the tumors via the cell cycle arrest, or the induced death via autophagy and apoptosis in the tumor cells, which is a type of programmed cell death.

The emission spectrum of the Chlorin E6 at the concentration of 5 × 10⁻⁸ M in the phosphate buffer solution at the pH of 7.4 is graphically presented in Figure 5. It is formed as the Chlorin E6 molecules return back to their original ground state.

Figure 3 The molecular basis of the mechanism of the Photodynamic therapy (PDT). The PS represents the Photosensitizer Compound that is Chlorin E6 in this case.

Figure 4 The Anti-tumor activity by the Chlorin E6, derivative of porphyrin. The produced ROS upregulate those genes that eventually contribute to the cell cycle arrest, autophagy and apoptosis

Figure 5 The Emission Spectrum of the Chlorin E6 is indicated by the continuous line. PBS stands for Phosphate buffer solution

The cells exhibiting either autophagy or apoptosis, release Damage-Associated Molecule Patterns (DAMPs) by which these cells are easily recognized by Antigen-Presenting Cells (APCs), which are the cells of body’s immune system. After being recognized, these cells/ tumors are presented to the CD8+ T cells. In this way, a systemic and a stronger immune response is initiated in the body against these cells/ tumors; that response is not only limited to the directly targeted cell but also against those tumor cells which are directly exposed to light. Immunogenic Cell Death (ICD) are the cells which are primarily involved in linking the killing of the cells directly exposed to light and the killing of those cells which are not exposed to the light. Thus, exhibiting a systemic immune response towards tumors. This mode of activity is known as abscopal effect. This concept of abscopal effect as a part of Photodynamic therapy (PDT) is presented in Figure 6. Interestingly, this abscopal effect has a clinical significance and a desirable event against metastases.

Figure 6 The Overview of the Photodynamic therapy (PDT) as a Cancer Therapy

In the murine model of melanoma and pancreatic tumor, abscopal antitumor effects is experimentally shown as the volume of the tumor is decreased in the right and also in the left side of the mouse body. The experimental results are graphically presented in Figure 7.

Figure 7 The Abscopal nature of the antitumor activity by the Chlorin E6. The effective Abscopal effect Abs_eff is indicated in red color graphs as compared to experimental control group

Chlorin E6 is used as a photosensitizer in the Photodynamic therapy (PDT) as it for treating melanoma as it is not only effective but also has a high tolerance. In addition, Chlorin E6 absorbs light of long wavelength, i.e., 660 nm; thus, thereby the penetration of Chlorin E6 in the biological tissue is also enhanced. Clinically, Chlorin E6 is a pharmaceutical drug which is also a photosensitizer. Chlorin E6 approved by FDA for the treatment of cancer via Photodynamic therapy (PDT). Although, in the research studies convincing results have been shown by Chlorin E6 against the skin cancer, pancreatic cancer, bladder cancer and breast cancer.

On the similar basis, Chlorin E6 is used as a sonosensitizer in the Sono-photodynamic therapy (SPDT) in the animal testing study. In that study, 4T1 mammary cancer cells of the mouse were used. Sono-photodynamic therapy (SPDT) is the treatment that combines both the Sonodynamic therapy (SDT) and the Photodynamic therapy (PDT). The cytotoxic/ antitumor effect of Chlorin E6 as a part of Photodynamic therapy (PDT) and Sonodynamic therapy (SDT) alone and also their combined effect, on the murine cells in in vitro laboratory conditions is graphically shown in Figure 8.

Figure 8 The Cytotoxic/ Antitumor effect of Chlorin E6. When the target cells were stimulated by light as well as sound, significant decrease in the number of living cells is achieved as compared to either of these stimuli independently