Drosophila and its uses

Abstract

A Drosophila containing xenotransplanted human cells; a method for evaluating a test substance, the method comprising the steps of administering a test substance to the Drosophila and evaluating the Drosophila; and a kit for evaluating a test substance, the kit comprising the Drosophila.

Description

[Technical field]

[0001] The present invention relates to Drosophila melanogaster and its use. More specifically, the present invention relates to Drosophila melanogaster, a method for evaluating a test substance, and a kit for evaluating a test substance. This application claims priority to US63 / 462,259, provisionally filed in the United States on April 27, 2023, the contents of which are incorporated herein by reference. [Background technology]

[0002] Nonclinical trials are essential in the rigorous development process of cancer therapeutics to ensure the safety and efficacy of drugs before human trials. Mouse models are the most commonly used animal models for nonclinical trials. Among them, patient-derived xenografts (PDX) models have been attracting attention in recent years.

[0003] In PDX models, a patient's tumor tissue is directly implanted into an immunodeficient mouse to mimic the patient's cancer. Drug testing is then performed to identify the most effective treatment for that case as a form of personalized medicine. However, xenograft models in which a patient's tumor tissue is implanted into a mouse are expensive, time-consuming to create, and require specialized facilities that are specific pathogen-free. In addition, large-scale screening is difficult due to ethical issues.

[0004] Therefore, it is desirable to establish simpler drug screening models that may lead to breakthroughs in the development of personalized medicine.

[0005] Meanwhile, Drosophila melanogaster has a short life span and a high reproductive rate, which allows large-scale research and screening at a lower cost than vertebrate models. Drosophila melanogaster has homologs of about 75% of human disease-related genes and also has many conserved signaling pathways. For this reason, it has been used as a model to understand the mechanisms of human diseases, including cancer.

[0006] For example, allograft tumor assays, in which larval tissue is injected into an adult host using the microinjection technique developed by Beadle and Ephrussi, are very useful for studying mechanisms of tumor-host interaction.

[0007] One example of a successful allograft model is the development of a mutant oncogene, Ras V12 Expression of Ras and loss of the tumor suppressor gene scribble induces Ras expression from the eye-antenna disc. V12 / scrib - / - After allografting into adult flies, V12 / scrib - / - Tumor tissues show extensive proliferation and infiltration into adjacent tissues (see, for example, Non-Patent Documents 1 and 2). [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] Figueroa-Clarevega A. and Bilder D., Malignant Drosophila tumors interrupt insulin signaling to induce cachexia-like wasting, Dev Cell, 33 (1), 47-55, 2015. [Non-Patent Document 2] Mishra-Gorur K., et al., Spz / Toll-6 signal guides organotropic metastasis in Drosophila, Dis Model Mech., 12 (10), dmm039727, 2019. Summary of the Invention [Problem to be solved by the invention]

[0009] However, despite many achievements in allograft models, no xenograft techniques have been reported in Drosophila. The present invention aims to provide a xenograft technique in Drosophila. [Means for solving the problem]

[0010] The present invention includes the following aspects. [1] Drosophila containing xenografted human cells. [2] An adult Drosophila described in [1]. [3] The Drosophila according to [1] or [2], wherein the human cell is a cancer cell. [4] The Drosophila according to [3], wherein the cancer cells are cells derived from a cancer patient. [5] A fruit fly according to any one of [1] to [4], which can be maintained at 28 to 40°C. [6] The Drosophila according to any one of [1] to [5], which is immunodeficient. [7] A method for evaluating a test substance, comprising the steps of administering the test substance to a fruit fly according to any one of [1] to [6], and evaluating the fruit fly. [8] The evaluation method described in [7], wherein the step of evaluating the Drosophila is a step of evaluating at least one selected from the group consisting of proliferation of the human cells, gene expression in the human cells, and survival rate of the Drosophila. [9] The evaluation method according to [7] or [8], wherein the evaluation method is for evaluating the anticancer effect of a test substance.

[10] A kit for evaluating a test substance, comprising the Drosophila melanogaster described in any one of [1] to [6]. Effect of the Invention

[0011] According to the present invention, a xenotransplantation technique in Drosophila can be provided. [Brief description of the drawings]

[0012] [Figure 1] FIG. 1 is a schematic diagram illustrating the experimental scheme of Experimental Example 1. [Diagram 2] FIG. 2 is an image showing the results of observing the fluorescence of tumor cells in the abdomen of a host fly in Experimental Example 1. [Diagram 3] FIG. 3 is an image showing the results of observing the fluorescence of tumor cells in the abdomen of a host fly in Experimental Example 1. [Figure 4] FIG. 4 is a graph showing the percentage of host flies in which fluorescence of tumor cells was observed in Experimental Example 1. [Diagram 5] FIG. 5 is a graph showing the number of days it took for the fluorescence of tumor cells in the host fly to disappear in Experimental Example 1. [Figure 6] FIG. 6 is a graph showing the survival rate of the host flies in Experimental Example 1. [Figure 7] FIG. 7 is a graph showing the results of measuring cell proliferation in Experimental Example 1. [Figure 8] FIG. 8 is an image showing the results of observing the fluorescence of tumor cells in the abdomen of a host fly in Experimental Example 2. [Figure 9] FIG. 9 shows images showing the results of observing the fluorescence of tumor cells in the abdomen of the host fly in Experimental Example 2. [Figure 10] FIG. 10 is an image showing the results of observing the fluorescence of A549 cells in the abdomen of the host fly in Experimental Example 3. [Figure 11] FIG. 11 is a graph showing the percentage of host flies in which fluorescence from A549 cells was observed in Experimental Example 3. [Figure 12] FIG. 12 is a graph showing the number of days it took for the fluorescence of A549 cells in the host fly to disappear in Experimental Example 3. [Figure 13] FIG. 13 is a graph showing the survival rate of each host fly in Experimental Example 3. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0013] [Drosophila] In one embodiment, the present invention provides a Drosophila containing xenotransplanted human cells. The Drosophila of this embodiment can be used as a xenotransplantation model and is useful for disease research, drug screening, and the like. Conventionally, Drosophila has not been used as a target for xenotransplantation of human cells due to differences in growth temperatures between Drosophila and human cells. In response to this, the inventors have demonstrated that Drosophila can be used as a xenotransplantation model, as described later in the Examples.

[0014] The Drosophila of this embodiment is preferably an adult. Drosophila becomes an adult in about 10 days from egg. Specifically, the egg hatches in about one day and becomes a first instar larva. The first instar larva molts and becomes a second instar larva about one day later. The second instar larva molts and becomes a third instar larva about one day later, and becomes a pupa about two days later. The pupa emerges and becomes an adult. As described later in the Examples, human cells can be transplanted into adult Drosophila and used as a xenotransplantation model.

[0015] The human cells transplanted into the Drosophila may be cancer cells. As described later in the Examples, Drosophila transplanted with human cancer cells can be suitably used for cancer research and drug screening.

[0016] Cancer cells to be transplanted into Drosophila may be cells derived from cancer patients. Drosophila transplanted with cancer cells from cancer patients can be used as a patient-derived xenograft (PDX) model. This model is less expensive and can be produced in a short time compared to mouse-based PDX models, and large-scale screening is also possible.

[0017] As described later in the Examples, the Drosophila of this embodiment can be preferably maintained at a temperature at which human cells are normally maintained. More specifically, it is preferable that the Drosophila can be maintained at 28 to 40°C, more preferably 29 to 38°C, and more preferably about 37°C. In this specification, "about" means a value including ±5%. For example, about 37°C can be said to be 35.15 to 38.85°C.

[0018] Drosophila are usually reared at a temperature of 14 to 29° C. As described later in the Examples, a xenograft model can be prepared by culturing Drosophila transplanted with human cells at 29° C.

[0019] In contrast, by using more thermotolerant Drosophila species, it may be possible to develop xenograft models that can be maintained at approximately 37°C. Some Drosophila species (e.g., Drosophila mojavensis) exhibit temperature tolerance and can be maintained at temperatures in the mid-30s to around 40°C.

[0020] The Drosophila of this embodiment is preferably immunodeficient. As described below in the Examples, transplanting human cells into an immunodeficient Drosophila allows the transplanted human cells to be maintained for a longer period of time.

[0021] Compared to vertebrates that have both adaptive and innate immunity, the immune system of Drosophila consists only of innate immunity regulated by the Toll pathway and the Imd pathway. Immunodeficient Drosophila includes mutants of any of the factors that compose these pathways. More specifically, for example, Dredd D44 , PGRP-LC Δ5 , Rel E20 , Tl 3 , Tl r3 , Tak1 2 , Myd88 c03881 , Myd88 kra-1 , spz 2 , spz 4 , spz 20 , IKKε1 , IKKε alice、 cact 3 , cact 9 , cact E8 Immunodeficient Drosophila mutants such as can be used.

[0022] [Test substance evaluation method] In one embodiment, the present invention provides a method for evaluating a test substance, the method comprising the steps of administering the test substance to a Drosophila containing xenografted human cells, and evaluating the Drosophila.

[0023] Drosophila containing xenografted human cells include those similar to those described above.

[0024] The test substance is not particularly limited, and examples thereof include natural compound libraries, synthetic compound libraries, existing drug libraries, metabolite libraries, etc. Methods for administering the test substance include oral administration.

[0025] Methods for evaluating Drosophila include methods for evaluating the proliferation of human cells xenografted into Drosophila, methods for evaluating gene expression in human cells, and methods for evaluating the survival rate of Drosophila.

[0026] The proliferation of human cells can be measured, for example, by fluorescently labeling human cells in advance and measuring the amount of fluorescence. Fluorescent labeling of human cells can be performed, for example, by expressing a fluorescent protein such as GFP, YFP, or mCherry in human cells. Evaluation of the survival rate of Drosophila may be, for example, by measuring the survival rate of Drosophila after a predetermined time from administration of a test substance, or by measuring the lifespan of Drosophila. The survival rate of Drosophila can be determined by counting the number of live flies and the number of dead flies.

[0027] The evaluation method of the present embodiment may be for evaluating the anticancer effect of a test substance when the human cells xenografted into Drosophila are cancer cells. The human cells may be cancer cells derived from a cancer patient. In this case, the evaluation method of the present embodiment can be said to be an evaluation method for personalized medicine.

[0028] [Test substance evaluation kit] In one embodiment, the present invention provides a kit for evaluating a test substance, comprising a Drosophila having xenografted human cells. The above-mentioned method for evaluating a test substance can be suitably carried out using the kit of this embodiment. Examples of the Drosophila having xenografted human cells include the same as those described above. EXAMPLES

[0029] The present invention will now be described in more detail with reference to examples, but the present invention is not limited to the following examples.

[0030] Materials and Methods (Drosophila strains and stocks) y, w, ey-Flp;Act>y+>Gal4, UAS-GFP;FRT82B, Tub-Gal80 and UAS-Ras V12 was a gift from T. Xu, Yale University School of Medicine. FRT82B and scrib / TM6B were gifts from D. Bilder, University of California, Berkeley. E20 (Stock number: 9457), Dredd D44 (Stock number: 80924), and PGRP-LC Δ5 (Stock number: 36323) was obtained from the Bloomington Drosophila Stock Center (Indiana, USA).

[0031] w 1118 were maintained at 25°C on standard medium. E20 , Dredd D44 , and PGRP-LC Δ5 were maintained at 20°C on standard medium.

[0032] (cell culture) Mammalian cell lines were maintained at 37°C and 5% CO2 in Dulbecco's modified Eagle's medium (DMEM, high glucose, Fujifilm Wako Pure Chemical Industries, catalog number: 044-29765) supplemented with 10% fetal bovine serum (FBS, Biocera, catalog number: 12389) and 100 μm / mL penicillin and 100 μg / mL streptomycin (Nacalai Tesque, catalog number: L5B3459).

[0033] HeLa Kyoto EGFP-α-tubulin / H2B-mCherry, A549 YFP-keratin 8, MCF7 YFP-keratin 8, and DLD-1 GFP-lifeact were kindly provided by K. Ohashi (Tohoku University).

[0034] (Ras V12 / scrib - / - Cloning and allogeneic transplantation Ras tagged with green fluorescent protein (GFP) V12 / scrib - / - Malignant tumor tissues were prepared in larval eye discs using the ey-FLP MARCM (eyMARCM) system. Subsequently, tumor tissues were dissected and collected from third instar larvae. The tumor tissues were then treated with 0.083% trypsin (Fujifilm Wako Pure Chemicals, Catalog No.: TWF7014) for 5 min, passed through a 27G needle (Terumo, Catalog No.: NN-2719S) using a 1 mL syringe, and finally suspended in phosphate-buffered saline (PBS, Fujifilm Wako Pure Chemicals). The cell suspension was then placed in a glass needle (Drummond Scientific Company, Catalog No.: 1-000-0300) and injected into the abdomen of young adult female flies using a microinjector (Narishige, IM300). The settings of the microinjector were N2 gas filling pressure: 20.0 PSI, injection pressure: 5.0 PSI, and balance pressure: 1.8 PSI. Injection of tumor tissue was performed within 5 days after excision. The host flies transplanted with tumor tissue were incubated at 29°C.

[0035] (implantation of human tumor cell lines) Human tumor cell lines, HeLa Kyoto EGFP-α-tubulin, A549 / YFP-keratin 8, MCF7 / YFP-keratin 8, and DLD-1 / GFP-lifeact, were treated with 0.05% to 0.083% trypsin. They were then pelleted, resuspended in 1 mL of PBS, and counted. After counting, the cells were suspended in fresh PBS at the desired concentration.

[0036] The cell concentration for injection was 600 x 10 for human tumor cells. 4 pieces / 50μL PBS, Ras V12 / scrib - / - About 300 x 10 4 (2) 300 x 10 cells for A549 cell injection into immunodeficient flies 4 The concentration was 50 μL / 50 μL PBS.

[0037] The cell suspension was injected into the abdomen of young adult female flies in the same manner as described above for allografts. 1118 , Rel E20 , Dredd D44 , or PGRP-LC Δ5 The host flies implanted with tumor cells were incubated at 29°C.

[0038] (Temperature-controlled cell proliferation assay) Human tumor cell lines, HeLa Kyoto EGFP-α-tubulin / H2B-mCherry, A549 / YFP-keratin 8, MCF7 / YFP-keratin 8, and DLD-1 / GFP-lifeact, were treated with 0.05%–0.083% trypsin, pelleted, and resuspended in 1 mL of PBS. Cells were then counted in a Burker-Turk Counting chamber (Sunlead glass) and plated at 5.0 × 10 cells per well in a 6-well plate. 4 The cells were then incubated at 37° C. for 3 days or at 29° C. for 7 days, during which the cells were counted daily.

[0039] (Measuring fruit fly life span) Flies were transferred to fresh food daily for 7 days after injection and every 2 days thereafter to avoid infection. Live flies were counted every 2 days. Life span analysis was performed and log-rank test statistics were calculated using R version 4.0.2 (R Foundation).

[0040] (histochemistry) Three days after injection, the host fly abdomen was dissected and the ventral epidermis was removed. The dissected abdomen was washed with PBS to remove floating cells in the hemolymph, leaving only cells that had interacted with the host tissue. The samples were then fixed with 4% paraformaldehyde (PFA) and washed three times with PBS. The cell nuclei were then stained with Hoechst33342 (Thermo Fisher Scientific, Catalog No.: H1399) and F-actin was stained with Alexa555 phalloidin (1:500, Thermo Fisher Scientific, Catalog No.: A34055). The samples were then washed with 0.1% TritonX-100 / PBS (PBST) and mounted on glass slides.

[0041] All imaging was performed using a confocal laser scanning microscope (Leica, TCS SP8) with either a 40x or 63x NA1.3 oil immersion objective (Leica). Image analysis was performed using the ImageJ software program (Open Source).

[0042] [Experimental Example 1] Ras V12 / scrib - / - Expression of ocular disc and Ras V12 Ectopic transplantation of tumorigenic tissues and cells, such as primary cell lines expressing IL-1, into adult flies has been reported to overgrow and promote the death of the host. To determine the behavior of human tumor cells in the fly's body, the inventors injected fluorescently labeled cultured human tumor cells into the abdomen of flies.

[0043] For xenografts, we used (1) HeLa-Kyoto (human cervical cancer cells), (2) A549 (lung epithelial adenocarcinoma cells), (3) MCF7 (breast cancer cells), and (4) DLD-1 (colorectal adenocarcinoma cells). As a positive control for tumor growth, Ras V12 / scrib - / - Cells were used.

[0044] Figure 1 is a schematic diagram illustrating the experimental scheme of this experimental example. After human tumor cells were transplanted into host flies, the host flies were incubated at 29°C, and the spread of the tumor was evaluated by monitoring the fluorescence of the tumor cells in the abdomen with a fluorescent microscope. The survival rate of the host flies was also examined.

[0045] Figures 2 and 3 are images showing the results of observing the fluorescence of tumor cells in the abdomen of a host fly. In Figures 2 and 3, the arrows indicate the fluorescence of tumor cells. Figure 4 is a graph showing the percentage of host flies in which fluorescence of tumor cells was observed. Figure 5 is a graph showing the number of days until the fluorescence of tumor cells in host flies disappeared. In Figure 5, "NS" indicates no significant difference, and "***" indicates a significant difference at p<0.001. Figure 6 is a graph showing the survival rate of each host fly. In Figure 6, "NS" indicates no significant difference, and "**" indicates a significant difference at p<0.01.

[0046] The fluorescence of HeLa and MCF7 cells disappeared 3-5 days after injection. In contrast, the fluorescence of A549 and DLD-1 cells persisted for 10-13 days. There were also differences in the patterning of fluorescence depending on the tumor cell line. A549 cells spread and persisted in a larger area of ​​the abdomen. DLD-1 cells frequently clustered and showed spot-like fluorescence within the host abdomen. Ras V12 / scrib - / - Cells showed progressive proliferation over a 7-day period, which was associated with very low viability.

[0047] Xenograft flies implanted with human tumor cells express Ras V12 / scrib - / -However, xenograft flies, except for those transplanted with HeLa cells, died significantly sooner than PBS-injected control flies.

[0048] Overall, the human tumor cells were able to survive in the host flies for several days before being eliminated. V12 / scrib - / - Cell-transplanted flies showed expanded fluorescence.

[0049] One potential factor influencing the reduced survival rate of human tumor cells in the fly's body was the optimal temperature difference between human and fly cells. The reported optimal temperature for healthy Drosophila melanogaster is 14-29°C, and the incubation temperature in the laboratory is 25°C. In contrast, mammalian cells are usually cultured at a much higher temperature of 37°C, and it was thought that they might undergo cellular stress when cultured at lower temperatures. Therefore, human tumor cell lines were cultured at 29°C and 37°C and cell proliferation was examined.

[0050] Figure 7 is a graph showing the results of measuring cell proliferation. As a result, the human tumor cell lines proliferated exponentially at different rates at 37°C, with HeLa cells and A549 cells showing the fastest proliferation rates.

[0051] When incubated at 29°C, most tumor cell lines showed a decrease in proliferation rate compared to 37°C. HeLa cells showed a decrease in number over time at 29°C. A549 cells still maintained proliferation at 29°C, albeit at a slower rate than at 37°C. These results indicate that different tumor cell lines have different proliferation rates at 37°C and 29°C. They also indicate that different tumor cell lines have different abilities to adapt to low temperatures.

[0052] Based on the results of xenograft and in vitro culture experiments, the inventors considered the correlation between the temperature tolerance of cell lines and their behavior after transplantation into host flies. A549 cells, which continued to grow even at 29°C, showed persistent GFP fluorescence until about day 13 when transplanted into flies, and induced a lower survival rate of host flies. Also, HeLa cells, which are less tolerant to low temperatures than other cells, could not survive for more than 3 days when transplanted into flies.

[0053] These results demonstrated that cell survival in xenograft flies differs depending on the cell type. Even when xenografting cells with different temperature ranges from the host fly, the method was shown to be able to maintain cell survival in xenograft flies for a long period of time and to be able to adequately evaluate the cell survival by appropriately selecting the cells.

[0054] Although both HeLa and MCF7 cells continued to survive in vitro at 29° C., when transplanted into flies, they disappeared from the bodies of Drosophila melanogaster within 5 days, so the cause of this was examined.

[0055] [Experimental Example 2] The results of measuring the life span of host flies transplanted with human tumor cells showed that the presence and behavior of human tumor cells had a deleterious effect on the host flies. Therefore, to investigate the interaction between human tumor cells and fly tissues at the cellular level, we closely observed fluorescently labeled human tumor cells 3 days after transplantation.

[0056] Prior to fluorescent staining of the host flies, the samples were washed with PBS to remove cells suspended in the hemolymph. After fixation and staining, the samples were imaged using a confocal laser microscope. A549 cells were selected as human tumor cells, as they persist longer in the host flies than other cell lines. For comparison, allografts were also performed in addition to xenografts of A549 cells. Two Drosophila tumor models, Ras V12 and Ras V12 / scrib - / - were observed as negative and positive controls for tumorigenicity, respectively.

[0057] 8 and 9 are fluorescence microscopy images. In FIG. 8, an enlarged image of the boxed area in b is shown in b', and an enlarged image of the boxed area in c is shown in c'. As a result, Ras V12 / scrib - / - The cells were V12 They grew better and formed larger clusters than benign tumors. A549 cells were sporadically located adjacent to the host fly tissue. As shown in Figure 9, A549 cells were able to express Ras despite having a reduced growth rate at 29°C compared to 37°C. V12 / scrib - / - They showed similar clustering behavior to A549 cells, forming distinct clusters within the muscle layer of the intestine. These results, together with the lifespan measurements, indicate that other factors play a role in preventing the survival of A549 cells in the host fly.

[0058] [Experimental Example 3] Drosophila, like many insects, is known to have only an innate immune system. Drosophila innate immunity consists of two pathways, the Toll pathway and the Imd pathway. Both of these pathways induce the production of antibacterial and antifungal peptides, including drosomycin and diptericin, to fight bacterial or fungal infections. However, it remains unclear how the Drosophila immune system responds to foreign cells, especially those from more phylogenetically distant organisms such as human cells.

[0059] The results that A549 cells were visibly adjacent to the host tissue 3 days after injection but gradually disappeared over time indicate that Drosophila innate immunity may be involved in the rejection of human tumor cells.

[0060] Therefore, the inventors have identified an Imd pathway component mutant, Dredd D44 , PGRP-LC Δ5 and Rel E20We transplanted human tumor cells into these mutant flies and examined their effects on the growth of A549 cells. These Drosophila strains are widely used because viable homozygous lines are readily available. We transplanted A549 cells into these mutant flies and examined the effects of innate immunity on the growth of A549 cells.

[0061] FIG. 10 is an image showing the results of observing the fluorescence of A549 cells in the abdomen of a host fly. In FIG. 10, the arrow indicates the fluorescence of A549 cells. FIG. 11 is a graph showing the percentage of host flies in which fluorescence of A549 cells was observed. FIG. 12 is a graph showing the number of days until the fluorescence of A549 cells in the host flies disappeared. In FIG. 12, "***" indicates a significant difference at p<0.001. FIG. 13 is a graph showing the survival rate of each host fly. In FIG. 13, "NS" indicates no significant difference, "*" indicates a significant difference at p<0.05, and "**" indicates a significant difference at p<0.01.

[0062] As a result, the transplanted A549 cells were compared with the control 1118 Compared with only an average of 10 days in flies, PGRP-LC Δ5 and Dredd D44 Hosts average 14-16 days, and Rel E20 It was shown that immune-deficient flies survived significantly longer before death for the host.

[0063] Furthermore, transplanted A549 cells showed 1118 The fluorescence was more widespread and more intense in immunodeficient flies than in nonimmunodeficient flies. 1118 Similar to injections in flies, the location of fluorescence was consistent over time for A549 cells in immunodeficient host flies.

[0064] These results indicate that A549 cells can survive longer in flies lacking a functional innate immune system. We then examined the effect of transplanting A549 cells into immunodeficient hosts by measuring the life span of the host flies. As a result, as shown in Figure 13, a significant early decrease in survival rate was observed immediately after transplantation of A549 cells.

[0065] Since the presence of A549 cells appears to be the primary cause of death in the host flies, which may be due to the stress associated with the injection, Figure 13 also shows measurements of the life span of uninjected and PBS-injected flies.

[0066] The results showed that PBS injection significantly increased the expression of PGRP-LC compared to non-injected flies. Δ5 Dredd causes a small but significant increase in fly mortality. D44 Given that injection of A549 cells caused a significant decrease in survival compared to controls, it appears that human tumor cells have a deleterious effect on the host fly.

[0067] These results indicate that although the immune system of the host fly is a significant obstacle to the survival of xenografted cells, the transplanted human tumor cells can survive and cause negative effects on the Drosophila host. Therefore, Drosophila xenografted with human tumor cells can be used for the evaluation of test substances. [Industrial Applicability]

[0068] According to the present invention, a xenotransplantation technique in Drosophila can be provided.

Claims

1. Drosophila containing xenografted human cells.

2. The Drosophila of claim 1, which is an adult.

3. The Drosophila according to claim 1 or 2, wherein the human cell is a cancer cell.

4. The Drosophila according to claim 3 , wherein the cancer cells are derived from a cancer patient.

5. The fruit fly according to claim 1 or 2, which can be maintained at 28 to 40°C.

6. The Drosophila of claim 1 or 2, which is immunodeficient.

7. A method for evaluating a test substance, comprising: A step of administering the test substance to the Drosophila according to claim 1 or 2; evaluating the Drosophila.

8. evaluating the Drosophila The evaluation method according to claim 7 , which comprises evaluating at least one selected from the group consisting of proliferation of the human cells, gene expression in the human cells, and survival rate of the Drosophila.

9. The evaluation method according to claim 7 , wherein the evaluation method is for evaluating an anticancer effect of a test substance.

10. A kit for evaluating a test substance, comprising the Drosophila according to claim 1 or 2.

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