Dorsomedial and preoptic hypothalamic circuits control torpor.

Endotherms can survive low temperatures and food shortage by actively entering a hypometabolic state known as torpor. Although the decrease in metabolic rate and body temperature (Tb) during torpor is controlled by the brain, the specific neural circuits underlying these processes have not been comprehensively elucidated. In this study, we identify the neural circuits involved in torpor regulation by combining whole-brain mapping of torpor-activated neurons, cell-type-specific manipulation of neural activity, and viral tracing-based circuit mapping. We find that Trpm2-positive neurons in the preoptic area and Vgat-positive neurons in the dorsal medial hypothalamus are activated during torpor. Genetic silencing shows that the activity of either cell type is necessary to enter the torpor state. Finally, we show that these cells receive projections from the arcuate and suprachiasmatic nucleus and send projections to brain regions involved in thermoregulation. Our results demonstrate an essential role of hypothalamic neurons in the regulation of Tb and metabolic rate during torpor and identify critical nodes of the torpor regulatory network.

Contrast-enhanced ultrasound imaging for monitoring the efficacy of near-infrared photoimmunotherapy.

BACKGROUND: Near-infrared photoimmunotherapy (NIR-PIT) is a promising cancer therapy combining NIR-light irradiation with an antibody and IR700DX, a light-sensitive substance, to destroy tumours. However, homogeneous irradiation is difficult because the light varies depending on the distance and tissue environment. Therefore, markers that indicate sufficient irradiation are necessary. Nanoparticles sized 10∼200 nm show enhanced permeation and retention within tumours, which is further enhanced via NIR-PIT (super enhanced permeability and retention, SUPR). We aimed to monitor the effectiveness of NIR-PIT by measuring SUPR. METHODS: A xenograft mouse tumour model was established by inoculating human cancer cells in both buttocks of Balb/C-nu/nu mice, and NIR-PIT was performed on only one side. To evaluate SUPR, fluorescent signal examination was performed using QD800-fluorescent nanoparticles and NIR-fluorescent poly (d,l-lactide-co-glycolic acid) (NIR-PLGA) microparticles. Harmonic signals were evaluated using micro-bubbles of the contrast agent Sonazoid and contrast-enhanced ultrasound (CEUS) imaging. The correlation between SUPR immediately after treatment and NIR-PIT effectiveness on the day after treatment was evaluated. FINDINGS: QD800 fluorescent signals persisted only in the treated tumours, and the intensity of remaining signals showed high positive correlation with the therapeutic effect. NIR-PLGA fluorescent signals and Sonazoid-derived harmonic signals remained for a longer time in the treated tumours than in the controls, and the kE value of the two-compartment model correlated with NIR-PIT effectiveness. INTERPRETATION: SUPR measurement using Sonazoid and CEUS imaging could be easily adapted for clinical use as a therapeutic image-based biomarker for monitoring and confirming of NIR-PIT efficacy. FUNDING: This research was supported by ARIM JAPAN of MEXT, the Program for Developing Next-generation Researchers (Japan Science and Technology Agency), KAKEN (18K15923, 21K07217) (JSPS), CREST (JPMJCR19H2, JST), and FOREST-Souhatsu (JST). Mochida Memorial Foundation for Medical and Pharmaceutical Research; Takeda Science Foundation; The Japan Health Foundation; and Princess Takamatsu Cancer Research Fund. Funders only provided financial support and had no role in the study design, data collection, data analysis, interpretation, and writing of the report.

Inside-the-body light delivery system using endovascular therapy-based light illumination technology.

BACKGROUND: Light-based therapies are promising for treating diseases including cancer, hereditary conditions, and protein-related disorders. However, systems, methods, and devices that deliver light deep inside the body are limited. This study aimed to develop an endovascular therapy-based light illumination technology (ET-BLIT), capable of providing deep light irradiation within the body. METHODS: The ET-BLIT system consists of a catheter with a single lumen as a guidewire and diffuser, with a transparent section at the distal end for thermocouple head attachment. The optical light diffuser alters the emission direction laterally, according to the optical fibre's nose-shape angle. If necessary, after delivering the catheter to the target position in the vessel, the diffuser is inserted into the catheter and placed in the transparent section in the direction of the target lesion. FINDINGS: ET-BLIT was tested in an animal model. The 690-nm near-infrared (NIR) light penetrated the walls of blood vessels to reach the liver and kidneys without causing temperature increase, vessel damage, or blood component alterations. NIR light transmittance from the diffuser to the detector within the organ or vessel was approximately 30% and 65% for the renal and hepatic arteries, respectively. INTERPRETATION: ET-BLIT can be potentially used in clinical photo-based medicine, as a far-out technology. ET-BLIT uses a familiar method that can access the whole body, as the basic procedure is comparable to that of endovascular therapy in terms of sequence and technique. Therefore, the use of the ET-BLIT system is promising for many light-based therapies that are currently in the research phase. FUNDING: Supported by Programme for Developing Next-generation Researchers (Japan Science and Technology Agency); JSPS KAKENHI (18K15923, 21K07217); JST-CREST (JPMJCR19H2); JST-FOREST-Souhatsu (JPMJFR2017); The Uehara Memorial Foundation; Yasuda Memorial Medical Foundation; Mochida Memorial Foundation for Medical and Pharmaceutical Research; Takeda Science Foundation; The Japan Health Foundation; Takahashi Industrial and Economic Research Foundation; AICHI Health Promotion Foundation; and Princess Takamatsu Cancer Research Fund.

Near-Infrared Photoimmunotherapy for Thoracic Cancers: A Translational Perspective.

The conventional treatment of thoracic tumors includes surgery, anticancer drugs, radiation, and cancer immunotherapy. Light therapy for thoracic tumors has long been used as an alternative; conventional light therapy also called photodynamic therapy (PDT) has been used mainly for early-stage lung cancer. Recently, near-infrared photoimmunotherapy (NIR-PIT), which is a completely different concept from conventional PDT, has been developed and approved in Japan for the treatment of recurrent and previously treated head and neck cancer because of its specificity and effectiveness. NIR-PIT can apply to any target by changing to different antigens. In recent years, it has become clear that various specific and promising targets are highly expressed in thoracic tumors. In combination with these various specific targets, NIR-PIT is expected to be an ideal therapeutic approach for thoracic tumors. Additionally, techniques are being developed to further develop NIR-PIT for clinical practice. In this review, NIR-PIT is introduced, and its potential therapeutic applications for thoracic cancers are described.

HER2 targeting near-infrared photoimmunotherapy for a CDDP-resistant small-cell lung cancer.

BACKGROUND: Human epidermal growth factor receptor 2 (HER2) is tyrosine kinase receptor that belongs to the ErbB family and is overexpressed on the membrane surface of various cancer cells, including small cell lung cancer (SCLC); however, no HER2 targeted therapy for SCLC have yet been established. Near-infrared photoimmunotherapy (NIR-PIT) is a novel cancer therapy based on photo-absorber, IRDye-700DX (IR700), -antibody conjugates, and near-infrared (NIR) light. METHODS: We used HER2-positive SCLC parental cell lines (SBC-3) and its chemoresistant cell lines, and examined therapeutic efficacy of HER2 targeting NIR-PIT using anti HER2 antibody trastuzumab. RESULTS: We found that HER2 expression was upregulated on chemoresistant cell lines, especially cisplatin-resistance (SBC-3/CDDP). In vitro, the rate of cell death increased with the amount of NIR-light irradiation, and it was significantly higher in SBC-3/CDDP than in SBC-3. In vivo, tumor growth was more suppressed in SBC-3/CDDP group than in SBC-3 group, and survival period tended to be prolonged. CONCLUSION: In this study, we demonstrated that HER2 targeting NIR-PIT using trastuzumab is promising therapy for HER2-positive SCLC, and is more effective when HER2 expression is upregulated due to CDDP resistance, suggesting that the HER2 expression level positively corelated with the efficacy of NIR-PIT.

Spatiotemporal depletion of tumor-associated immune checkpoint PD-L1 with near-infrared photoimmunotherapy promotes antitumor immunity.

BACKGROUND: Near-infrared photoimmunotherapy (NIR-PIT) is a new modality for treating cancer, which uses antibody-photoabsorber (IRDye700DX) conjugates that specifically bind to target tumor cells. This conjugate is then photoactivated by NIR light, inducing rapid necrotic cell death. NIR-PIT needs a highly expressed targeting antigen on the cells because of its reliance on antibodies. However, using antibodies limits this useful technology to only those patients whose tumors express high levels of a specific antigen. Thus, to propose an alternative strategy, we modified this phototechnology to augment the anticancer immune system by targeting the almost low-expressed immune checkpoint molecules on tumor cells. METHODS: We used programmed death-ligand 1 (PD-L1), an immune checkpoint molecule, as the target for NIR-PIT. Although the expression of PD-L1 on tumor cells is usually low, PD-L1 is almost expressed on tumor cells. Intratumoral depletion with PD-L1-targeted NIR-PIT was tested in mouse syngeneic tumor models. RESULTS: Although PD-L1-targeted NIR-PIT showed limited effect on tumor cells in vitro, the therapy induced sufficient antitumor effects in vivo, which were thought to be mediated by the 'photoimmuno' effect and antitumor immunity augmentation. Moreover, PD-L1-targeted NIR-PIT induced antitumor effect on non-NIR light-irradiated tumors. CONCLUSIONS: Local PD-L1-targeted NIR-PIT enhanced the antitumor immune reaction through a direct photonecrotic effect, thereby providing an alternative approach to targeted cancer immunotherapy and expanding the scope of cancer therapeutics.

Hurdles for the wide implementation of photoimmunotherapy.

Near infrared photoimmunotherapy (NIR-PIT) is a molecularly targeted treatment for cancers achieved by injecting a conjugate of IRDye700DX® (IR700), a water-soluble silicon phthalocyanine derivative in the near infrared, and a monoclonal antibody that targets cancer cell antigens. NIR-PIT is a highly specific treatment with few side effects that results in rapid immunogenic cell death. Despite it being a very effective and innovative therapy, there are a few challenges preventing full implementation in clinical practice. These include the limits of near infrared light penetration, selection of targets, concerns about tumor lysis syndrome and drug costs. However, NIR-PIT has been approved by the regulatory authorities in Japan, allowing for exploration of how to mitigate challenges while maximizing the benefits of this treatment modality.

Near-infrared photoimmunotherapy targeting GPR87: Development of a humanised anti-GPR87 mAb and therapeutic efficacy on a lung cancer mouse model.

BACKGROUND: GPR87 is a G-protein receptor that is specifically expressed in tumour cells, such as lung cancer, and rarely expressed in normal cells. GPR87 is a promising target for cancer therapy, but its ligand is controversial. Near-infrared photoimmunotherapy (NIR-PIT) is a novel cancer therapy in which a photosensitiser, IRDye700DX (IR700), binds to antibodies and specifically destroys target cells by irradiating them with near-infrared-light. Here, we aimed to develop a NIR-PIT targeting GPR87. METHODS: We evaluated the expression of GPR87 in resected specimens of lung cancer and malignant pleural mesothelioma (MPM) resected at Nagoya University Hospital using immunostaining. Humanised anti-GPR87 antibody (huGPR87) was generated by introducing CDRs from mouse anti-GPR87 antibody generated by standard hybridoma method. HuGPR87 was conjugated with IR700 and the therapeutic effect of NIR-PIT was evaluated in vitro and in vivo using lung cancer or MPM cell lines. FINDINGS: Among the surgical specimens, 54% of lung cancer and 100% of MPM showed high expression of GPR87. It showed therapeutic effects on lung cancer and MPM cell lines in vitro, and showed therapeutic effects in multiple models in vivo. INTERPRETATION: These results suggest that NIR-PIT targeting GPR87 is a promising therapeutic approach for the treatment of thoracic cancer. FUNDING: This research was supported by the Program for Developing Next-generation Researchers (Japan Science and Technology Agency), KAKEN (18K15923, 21K07217, JSPS), FOREST-Souhatsu, CREST (JST).

Near Infrared Photoimmunotherapy for Mouse Models of Pleural Dissemination.

The efficacy of photoimmunotherapy can be evaluated more accurately with an orthotopic mouse model than with a subcutaneous one. A pleural dissemination model can be used for the evaluation of treatment methods for intrathoracic diseases such as lung cancer or malignant pleural mesothelioma. Near-infrared photoimmunotherapy (NIR-PIT) is a recently developed cancer treatment strategy that combines the specificity of tumor-targeting antibodies with toxicity caused by a photoabsorber (IR700Dye) after exposure to NIR light. The efficacy of NIR-PIT has been reported using various antibodies; however, only a few reports have shown the therapeutic effect of this strategy in an orthotopic model. In the present study, we demonstrate an example of efficacy evaluation of the pleural disseminated lung cancer model, which was treated using NIR-PIT.

Targeted Phototherapy for Malignant Pleural Mesothelioma: Near-Infrared Photoimmunotherapy Targeting Podoplanin.

Malignant pleural mesothelioma (MPM) has extremely limited treatment despite a poor prognosis. Moreover, molecular targeted therapy for MPM has not yet been implemented; thus, a new targeted therapy is highly desirable. Near-infrared photoimmunotherapy (NIR-PIT) is a recently developed cancer therapy that combines the specificity of antibodies for targeting tumors with toxicity induced by the photoabsorber after exposure to NIR-light. In this study, we developed a new phototherapy targeting podoplanin (PDPN) for MPM with the use of both NIR-PIT and an anti-PDPN antibody, NZ-1. An antibody-photosensitizer conjugate consisting of NZ-1 and phthalocyanine dye was synthesized. In vitro NIR-PIT-induced cytotoxicity was measured with both dead cell staining and luciferase activity on various MPM cell lines. In vivo NIR-PIT was examined in both the flank tumor and orthotopic mouse model with in vivo real-time imaging. In vitro NIR-PIT-induced cytotoxicity was NIR-light dose dependent. In vivo NIR-PIT led to significant reduction in both tumor volume and luciferase activity in a flank model (p < 0.05, NIR-PIT group versus NZ-1-IR700 group). The PDPN-targeted NIR-PIT resulted in a significant antitumor effect in an MPM orthotopic mouse model (p < 0.05, NIR-PIT group versus NZ-1-IR700 group). This study suggests that PDPN-targeted NIR-PIT could be a new promising treatment for MPM.

Near infrared photoimmunotherapy targeting DLL3 for small cell lung cancer.

BACKGROUND: Small cell lung cancer (SCLC) has a poor prognosis, and its treatment options are limited. Delta-like protein 3 (DLL3) is expressed specifically in SCLC and is considered a promising therapeutic target for patients with this disease. Rovalpituzumab tesirine (Rova-T) was the first antibody-drug conjugate targeting DLL3. Although Rova-T development was unfortunately terminated, DLL3 remains an ideal target for SCLC. Near infrared photoimmunotherapy (NIR-PIT) is a new form of cancer treatment that employs an antibody-photosensitiser conjugate followed by NIR light exposure and damage target cells specifically. In this study, we demonstrate DLL3-targeted NIR-PIT to develop a novel molecularly targeted treatment for SCLC. METHODS: The anti-DLL3 monoclonal antibody rovalpituzumab was conjugated to an IR700 photosensitiser (termed 'rova-IR700'). SCLC cells overexpressing DLL3 as well as non-DLL3-expressing controls were incubated with rova-IR700 and then exposed to NIR-light. Next, mice with SCLC xenografts were injected with rova-IR700 and irradiated with NIR-light. FINDINGS: DLL3-overexpressing cells underwent immediate destruction upon NIR-light exposure, whereas the control cells remained intact. The xenograft in mice treated with rova-IR700 and NIR-light shrank markedly, whereas neither rova-IR700 injection nor NIR-light irradiation alone affected tumour size. INTERPRETATION: Our data suggest that targeting of DLL3 using NIR-PIT could be a novel and promising treatment for SCLC. FUNDING: Research supported by grants from the Program for Developing Next-generation Researchers (Japan Science and Technology Agency), KAKEN (18K15923, JSPS), Medical Research Encouragement Prize of The Japan Medical Association, The Nitto Foundation, Kanae Foundation for the Promotion of Medical Science.

Implantable wireless powered light emitting diode (LED) for near-infrared photoimmunotherapy: device development and experimental assessment in vitro and in vivo.

PURPOSE: The aim of this study was to develop and assess a novel implantable, wireless-powered, light-emitting diode (LED) for near-infrared photoimmunotherapy (NIR-PIT). NIR-PIT is a recently developed cancer therapy that uses NIR light and antibody-photosensitizer conjugates and is able to induce cancer-specific cell death. Due to limited light penetration depth it is currently unable to treat tumors in deep tissues. Use of implanted LED might potentially overcome this limitation. RESULTS: The wireless LED system was able to emit NIR light up to a distance of 20 cm from the transmitter coil by using low magnetic fields as compliant with limits for use in humans. Results indicated that the LED system was able to kill tumor cells in vitro and to suppress tumor growth in implanted tumor-bearing mice. CONCLUSIONS: Results indicated that the proposed implantable wireless LED system was able to suppress tumor growth in vivo. These results are encouraging as wireless LED systems such as the one here developed might be a possible solution to treat tumors in deep regions in humans. Further research in this area would be important. MATERIALS AND METHODS: An implantable LED system was developed. It consisted of a LED capsule including two LED sources and a receiver coil coupled with an external coil and power source. Wireless power transmission was guaranteed by using electromagnetic induction. The system was tested in vitro by using EGFR-expressing cells and HER2-expressing cells. The system was also tested in vivo in tumor-bearing mice.

3D mesoscopic fluorescence tomography for imaging micro-distribution of antibody-photon absorber conjugates during near infrared photoimmunotherapy in vivo.

As a novel low-side-effect cancer therapy, photo-immunotherapy (PIT) is based on conjugating monoclonal antibody (mAb) with a near-infrared (NIR) phthalocyanine dye IRDye700DX (IR 700). IR700 is not only fluorescent to be used as an imaging agent, but also phototoxic. When illuminating with NIR light, PIT can induce highly-selective cancer cell death while leaving most of tumor blood vessels unharmed, leading to an effect termed super-enhanced permeability and retention (SUPR), which can significantly improve the effectiveness of anti-cancer drug. Currently, the therapeutic effects of PIT are monitored using 2D macroscopic fluorescence reflectance imager, which lacks the resolution and depth information to reveal the 3D distribution of mAb-IR700. In the study, we applied a multi-modal optical imaging approach including high-resolution optical coherence tomography (OCT) and high-sensitivity fluorescence laminar optical tomography (FLOT), to provide 3D tumor micro-structure and micro-distribution of mAb-IR700 in the tumor simultaneously during PIT in situ and in vivo. The multi-wavelength FLOT can also provide the blood vessels morphology of the tumor. Thus, the 3D FLOT reconstructed images allow us to evaluate the IR700 fluorescence distribution change with respect to the blood vessels and at different tumor locations/depths non-invasively, thereby enabling evaluation of the therapeutic effects in vivo and optimization of treatment regimens accordingly. The mAb-IR700 can access more tumor areas after PIT treatment, which can be explained by increased vascular permeability immediately after NIR-PIT. Two-photon microscopy was also used to record the mAb-IR700 on the tumor surface near the blood vessels to verify the results.

Near Infrared Photoimmunotherapy with Combined Exposure of External and Interstitial Light Sources.

Near infrared photoimmunotherapy (NIR-PIT) is a new target-cell-specific cancer treatment that induces highly selective necrotic/immunogenic cell death after systemic administration of a photoabsorber antibody conjugate and subsequent NIR light exposure. However, the depth of NIR light penetration in tissue (approximately 2 cm) with external light sources limits the therapeutic effects of NIR-PIT. Interstitial light exposure using cylindrical diffusing optical fibers can overcome this limitation. The purpose in this study was to compare three NIR light delivery methods for treating tumors with NIR-PIT using a NIR laser system at an identical light energy; external exposure alone, interstitial exposure alone, and the combination. Panitumumab conjugated with the photoabsorber IRDye-700DX (pan-IR700) was intravenously administered to mice with A431-luc xenografts which are epithelial growth factor receptor (EGFR) positive. One and 2 days later, NIR light was administered to the tumors using one of three methods. Interstitial exposure alone and in combination with external sources showed the greatest decrease in bioluminescence signal intensity. Additionally, the combination of external and interstitial NIR light exposure showed significantly greater tumor size reduction and prolonged survival after NIR-PIT compared to external exposure alone. This result suggested that the combination of external and interstitial NIR light exposure was more effective than externally applied light alone. Although external exposure is the least invasive means of delivering light, the combination of external and interstitial exposures produces superior therapeutic efficacy in tumors greater than 2 cm in depth from the tissue surface.

Evaluation of Early Therapeutic Effects after Near-Infrared Photoimmunotherapy (NIR-PIT) Using Luciferase-Luciferin Photon-Counting and Fluorescence Imaging.

Near-infrared photoimmunotherapy (NIR-PIT) is a newly developed cancer treatment that induces highly selective immunogenic cell death. It is based on an antibody-photoabsorber conjugate (APC) that is activated by NIR light. The purpose of this study was to investigate the effects of NIR-PIT as measured by luciferase-luciferin photon-counting and fluorescence imaging. Six days after subcutaneous injection of A431-luc-GFP cells tumors formed in a xenograft mouse model. The EGFR-targeting antibody, panitumumab, was conjugated to the photoabsorber, IRDye-700DX (pan-IR700), and was intravenously administered to tumor-bearing mice. Serial luciferase-luciferin photon-counting images and both green fluorescent protein (GFP) and IR700 fluorescence images were obtained from the same mice before and after NIR-PIT treatment (0, 10, 20, 30 min (early phase), and 24, 48 h (late phase) after NIR light exposure). Optical signal intensities were compared for each modality. IR700 fluorescence and luciferase-luciferin photon-counting images showed decreased intensities in both the early and late phases after NIR-PIT (p < 0.01). On the other hand, GFP fluorescence images showed decreased intensities only in the late phase (p < 0.01). In the early phase, GFP fluorescence images showed smaller intensity reductions compared to IR700 fluorescence and luciferase-luciferin photon-counting (p < 0.01), while in the late phase, IR700 fluorescence showed smaller intensity reductions than luciferase-luciferin photon-counting and GFP fluorescence (p < 0.05), due to redistribution of pan-IR700 within the tumor bed. In conclusion, luciferase-luciferin photon-counting imaging is suitable to evaluate early phase NIR-PIT effects, while both luciferase-luciferin photon-counting and GFP reflected later phase effects.

Real-time monitoring of microdistribution of antibody-photon absorber conjugates during photoimmunotherapy in vivo.

Photoimmunotherapy (PIT) is an emerging low side effect cancer therapy based on a monoclonal antibody (mAb) conjugated with a near-infrared (NIR) phthalocyanine dye IRDye 700DX. IR700 is fluorescent, can be used as an imaging agent, and also is phototoxic. It induces rapid cell death after exposure to NIR light. PIT induces highly selective cancer cell death, while leaving most of tumor blood vessels unharmed, leading to an effect called super-enhanced permeability and retention (SUPR). SUPR significantly improves the effectiveness of the anticancer drug. Currently, the therapeutic effects of PIT are monitored using the IR700 fluorescent signal based on macroscopic fluorescence reflectance imagery. This technique, however, lacks the resolution and depth information to reveal the intratumor heterogeneity of mAb-IR700 distribution. We applied a minimally invasive two-channel fluorescence fiber imaging system by combining the traditional fluorescence imaging microscope with two imaging fiber bundles (~0.85mm). This method monitored mAb-IR700 distribution and therapeutic effects during PIT at different intratumor locations (e.g., tumor surface vs. deep tumor) in situ and in real time simultaneously. This enabled evaluation of the therapeutic effects in vivo and treatment regimens. The average IR700 fluorescence intensity recovery after PIT to the tumor surface is 91.50%, while it is 100.63% in deep tumors. To verify the results, two-photon microscopy combined with a microprism was also used to record the mAb-IR700 distribution and fluorescence intensity of green fluorescent protein (GFP) at different tumor depths during PIT. After PIT treatment, there was significantly higher IR700 fluorescence recovery in deep tumor than in the tumor surface. This phenomenon can be explained by increased vascular permeability immediately after NIR-PIT. Fluorescence intensity of GFP at the tumor surface decreased significantly more compared to that of deep tumor and in controls (no PIT).

Cerenkov Radiation-Induced Photoimmunotherapy with 18F-FDG.

Near-infrared photoimmunotherapy (NIR-PIT) is a new cancer treatment that combines the specificity of antibodies for targeting tumors with toxicity induced by photoabsorbers after irradiation with NIR light. A limitation of NIR-PIT is the inability to deliver NIR light to a tumor located deep inside the body. Cerenkov radiation (CR) is the ultraviolet and blue light that is produced by a charged particle traveling through a dielectric medium faster than the speed of light in that medium and is commonly produced during radioactive decay. Here, we demonstrate the feasibility of using CR generated by 18F-FDG accumulated in tumors to induce photoimmunotherapy. Methods: Using A431-luc cells, we evaluated the therapeutic effects of CR-PIT in vitro and in vivo using bioluminescence imaging. Results: CR-PIT showed significant suppression of tumor size, but the decrease of bioluminescence after CR-PIT was not observed consistently over the entire time course. Conclusion: Although CR-PIT can induce tumor killing deep within body, it is less effective than NIR-PIT, possibly related to the relatively lower efficiency of short wavelength light than NIR.

Near Infrared Photoimmunotherapy in a Transgenic Mouse Model of Spontaneous Epidermal Growth Factor Receptor (EGFR)-expressing Lung Cancer.

Near infrared photoimmunotherapy (NIR-PIT) is a new cancer treatment that combines the specificity of antibodies for targeting tumors with the toxicity induced by a sensitive photoabsorber following exposure to NIR light. Most studies of NIR-PIT have been performed in xenograft models of cancer. The purpose of this study was to evaluate the therapeutic effects of NIR-PIT in a transgenic model of spontaneous lung cancer expressing human EGFR (hEGFR-TL). Mice were separated into 3 groups for the following treatments: (1) no treatment (control); (2) 150 µg of photoabsorber, IR700, conjugated to panitumumab, an antibody targeting EGFR [antibody-photoabsorber conjugate (APC)] intravenously (i.v.) only; (3) 150 µg of APC i.v. with NIR light administration. Each treatment was performed every week up to three weeks. MRI was performed 1 day before and 3, 6, 13, 20, 27, and 34 days after first NIR-PIT. The relative volume of lung tumors was calculated from the tumor volume at each MRI time point divided by the initial volume. Steel test for multiple comparisons was used to compare the tumor volume ratio with that of control. Tumor volume ratio was inhibited significantly in the NIR-PIT group compared with control group (P < 0.01 at all time points). In conclusion, NIR-PIT effectively treated a spontaneous lung cancer in a hEGFR-TL transgenic mouse model. MRI successfully monitored the therapeutic effects of NIR-PIT. Mol Cancer Ther; 16(2); 408-14. ©2016 AACR.

Immunogenic cancer cell death selectively induced by near infrared photoimmunotherapy initiates host tumor immunity.

Immunogenic cell death (ICD) is a form of cell death that activates an adaptive immune response against dead-cell-associated antigens. Cancer cells killed via ICD can elicit antitumor immunity. ICD is efficiently induced by near-infrared photo-immunotherapy (NIR-PIT) that selectively kills target-cells on which antibody-photoabsorber conjugates bind and are activated by NIR light exposure. Advanced live cell microscopies showed that NIR-PIT caused rapid and irreversible damage to the cell membrane function leading to swelling and bursting, releasing intracellular components due to the influx of water into the cell. The process also induces relocation of ICD bio markers including calreticulin, Hsp70 and Hsp90 to the cell surface and the rapid release of immunogenic signals including ATP and HMGB1 followed by maturation of immature dendritic cells. Thus, NIR-PIT is a therapy that kills tumor cells by ICD, eliciting a host immune response against tumor.

Near infrared photoimmunotherapy with avelumab, an anti-programmed death-ligand 1 (PD-L1) antibody.

Near Infrared-Photoimmunotherapy (NIR-PIT) is a highly selective tumor treatment that employs an antibody-photo-absorber conjugate (APC). Programmed cell death protein-1 ligand (PD-L1) is emerging as a molecular target. Here, we describe the efficacy of NIR-PIT, using fully human IgG1 anti-PD-L1 monoclonal antibody (mAb), avelumab, conjugated to the photo-absorber, IR700DX, in a PD-L1 expressing H441 cell line, papillary adenocarcinoma of lung. Avelumab-IR700 showed specific binding and cell-specific killing was observed after exposure of the cells to NIR in vitro. In the in vivo study, avelumab-IR700 showed high tumor accumulation and high tumor-background ratio. Tumor-bearing mice were separated into 4 groups: (1) no treatment; (2) 100 µg of avelumab-IR700 i.v.; (3) NIR light exposure only, NIR light was administered; (4) 100 µg of avelumab-IR700 i.v., NIR light was administered. Tumor growth was significantly inhibited by NIR-PIT treatment compared with the other groups (p < 0.001), and significantly prolonged survival was achieved (p < 0.01 vs other groups). In conclusion, the anti-PD-L1 antibody, avelumab, is suitable as an APC for NIR-PIT. Furthermore, NIR-PIT with avelumab-IR700 is a promising candidate of the treatment of PD-L1-expressing tumors that could be readily translated to humans.