JP5224325B2 - B cell malignant lymphoma treatment - Google Patents
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Description
本発明は、B細胞悪性リンパ腫を治療するための薬剤に関するものである。 The present invention relates to a drug for treating B cell malignant lymphoma.
各国、特に先進国において悪性腫瘍は死亡原因の上位を占めているが、その根本的な治療法はいまだ確立していないのが現状である。悪性腫瘍の治療方法としては、主として外科的手術、放射線治療、および抗腫瘍薬の投与といった化学療法が挙げられ、それぞれに一長一短がある。 Although malignant tumors occupy the top causes of death in various countries, especially developed countries, the current cure has not been established yet. Methods for treating malignant tumors mainly include chemotherapy, such as surgery, radiation therapy, and administration of antitumor drugs, each having its advantages and disadvantages.
悪性腫瘍の中でも悪性リンパ腫はリンパ系に生じ、上皮細胞由来の癌などとは異なり、リンパ系自体が全身組織であることから外科的手術による切除は不可能である。よって、治療手段としては一般的に放射線療法と化学療法が採用される。 Among malignant tumors, malignant lymphoma occurs in the lymphatic system, and unlike the cancer derived from epithelial cells, the lymphatic system itself is a systemic tissue and cannot be removed by surgical operation. Therefore, radiation therapy and chemotherapy are generally employed as treatment means.
化学療法は全身に作用させることができるので、微小腫瘍組織や転移能を有する進行性悪性腫瘍に効果を示す。腫瘍細胞はもともと患者自身の細胞が正常な制御に反して無秩序に増殖したものであるので、放射線や抗腫瘍薬は腫瘍に対して治療効果を示すが、正常な細胞にもダメージを与えるという欠点がある。そこで、腫瘍に対する治療効果に優れ且つ副作用が少ないといった、より良い抗腫瘍薬の登場が期待されている。 Since chemotherapy can be applied to the whole body, it is effective for progressive tumors with microtumor tissue or metastatic potential. Since tumor cells are originally cells that proliferate in a disorderly manner against normal control, radiation and antitumor drugs have a therapeutic effect on tumors, but they also have the drawback of damaging normal cells There is. Therefore, the advent of better antitumor drugs that have excellent therapeutic effects on tumors and few side effects is expected.
しかし、悪性リンパ腫は全身に発生するという性質上、腫瘍細胞が完全に消失したことを証明することはできない。それどころか、いったん治療した後にも再発する例が多いため、治療が困難であると考えられる。 However, due to the nature of malignant lymphoma occurring throughout the body, it is not possible to prove that the tumor cells have completely disappeared. On the contrary, it is considered difficult to treat because there are many cases of recurrence after treatment.
悪性リンパ腫は様々な分類がされており、その発生比率は人種、国や地域などにより異なる。例えば、リンパ球にはB細胞、T細胞とNK細胞があり、我が国では70〜80%がB細胞悪性リンパ腫、20〜30%がT細胞悪性リンパ腫で、NK細胞悪性リンパ腫は5%以下に過ぎないというデータがある。この様に、B細胞悪性リンパ腫は悪性リンパ腫の中でも高い比率で発生することがあるので、有効な治療手段が求められている。 Malignant lymphomas are classified into various categories, and the incidence varies depending on race, country and region. For example, lymphocytes include B cells, T cells, and NK cells. In Japan, 70 to 80% are B cell malignant lymphomas, 20 to 30% are T cell malignant lymphomas, and NK cell malignant lymphomas are less than 5%. There is no data. Thus, since B cell malignant lymphoma may occur at a high ratio among malignant lymphomas, an effective therapeutic means is required.
ところで、HMGB1(High Mobility Group box 1)は、げっ歯類からヒトまで95%以上のアミノ酸配列が等しいタンパク質である。このHMGB1は正常細胞にも存在するが、敗血症(全身性炎症反応症候群)において放出される菌体内毒素であるLPS(リポ多糖)による刺激によって血中濃度が上昇し、最終的な組織障害をもたらす。よって、特許文献1記載の技術では、炎症性サイトカインカスケードの活性化を特徴とする症状を治療するためにHMGB1アンタゴニストを投与しており、治療対象としてリンパ腫も例示されている。しかし当該特許文献1には、致死量のLPSを投与した敗血症モデルマウスの死亡率が抗HMGB1抗体の投与により低下した実験例が開示されているものの、他の疾患に対する抗HMGB1抗体の効果は実証されていない。
By the way, HMGB1 (High Mobility Group box 1) is a protein having the same amino acid sequence of 95% or more from rodents to humans. Although this HMGB1 is also present in normal cells, the blood concentration is increased by stimulation with LPS (lipopolysaccharide) which is an endotoxin released in sepsis (systemic inflammatory response syndrome), resulting in final tissue damage . Therefore, in the technique described in
特許文献2には、壊死組織により誘導される副作用の治療のための組成物として、HMGB1抗体等を含む組成物が開示されている。しかし、当該副作用としては、近傍の生存細胞の活性化、骨髄細胞の動員および活性化、内皮のバリア機能の喪失、浮腫のみが例示されており、B細胞悪性リンパ腫については記載されていない。
また、本発明者らは従前より抗HMGB1抗体について研究を継続しており、特許文献3と4の通り、これまで抗HMGB1抗体の脳梗塞と脳血管攣縮に対する抑制効果を見出している。
上述した様に、抗腫瘍薬は一般的に腫瘍細胞のみならず正常細胞にもダメージを与え得るものである。また、B細胞悪性リンパ腫は悪性リンパ腫の中でも発生比率が比較的高いものであるので、より良い治療手段が求められている。 As described above, antitumor drugs can generally damage not only tumor cells but also normal cells. In addition, since B cell malignant lymphoma has a relatively high incidence rate among malignant lymphomas, a better therapeutic means is required.
そこで、本発明が解決すべき課題は、B細胞悪性リンパ腫に対する治療効果を有し且つ副作用の少ない薬剤を提供することにある。 Therefore, the problem to be solved by the present invention is to provide a drug having a therapeutic effect on B-cell malignant lymphoma and having few side effects.
本発明者らは、上記課題を解決すべく、B細胞悪性リンパ腫の治療に有効な薬剤につき種々検討を進めた。その結果、抗HMGB1モノクローナル抗体が非常に優れた効果を有することを見出して、本発明を完成した。 In order to solve the above problems, the present inventors have made various studies on drugs effective for the treatment of B-cell malignant lymphoma. As a result, it was found that the anti-HMGB1 monoclonal antibody has a very excellent effect, and the present invention was completed.
本発明のB細胞悪性リンパ腫治療薬は、抗HMGB1モノクローナル抗体を有効成分とすることを特徴とする。 The therapeutic agent for B-cell malignant lymphoma of the present invention comprises an anti-HMGB1 monoclonal antibody as an active ingredient.
抗HMGB1モノクローナル抗体の投与量としては、後述する実験結果より、1回当たり0.2〜5mg/(kg体重)が好適であると考えられる。また、本発明のB細胞悪性リンパ腫治療薬の主成分は抗体であることから、投与形態としては静脈投与が好適である。 As a dose of the anti-HMGB1 monoclonal antibody, it is considered that 0.2 to 5 mg / (kg body weight) per dose is suitable from the experimental results described later. In addition, since the main component of the therapeutic agent for B cell malignant lymphoma of the present invention is an antibody, intravenous administration is suitable as an administration form.
本発明の治療薬は、悪性リンパ腫の中でも比較的高い発生比率を示すB細胞悪性リンパ腫を効果的に抑制することができる。また、現在使用されている抗体薬剤を考慮すれば、重篤な副作用を生じる可能性は極めて少ないと考えられる。従って、本発明のB細胞悪性リンパ腫治療薬は、より良い治療手段が切望されているB細胞悪性リンパ腫を副作用なく効果的に治療できるものとして、極めて有用である。 The therapeutic agent of the present invention can effectively suppress B cell malignant lymphoma having a relatively high incidence among malignant lymphomas. In addition, considering the currently used antibody drugs, it is very unlikely that serious side effects will occur. Therefore, the therapeutic agent for B-cell malignant lymphoma of the present invention is extremely useful as an agent that can effectively treat B-cell malignant lymphoma for which better treatment means are desired without side effects.
本発明のB細胞悪性リンパ腫は、抗HMGB1モノクローナル抗体を有効成分とする。 The B cell malignant lymphoma of the present invention contains an anti-HMGB1 monoclonal antibody as an active ingredient.
B細胞悪性リンパ腫は非ホジキン悪性リンパ腫の一種である。非ホジキン悪性リンパ腫にはB細胞悪性リンパ腫とT細胞悪性リンパ腫があるが、人種や地域などによってはT細胞悪性リンパ腫よりもB細胞悪性リンパ腫の発生比率の方が圧倒的に高い場合がある。その症状としてはリンパ節の腫大が見られるが、一般的には無痛であるために自覚症状がないままに悪化することから問題である。B細胞悪性リンパ腫は全身性であることから、その治療手段としては、患者の年齢や重篤度などにもよるが、一般的には多剤併用化学療法が採用される。よって、抗がん剤の副作用による患者への負担が大きいことから、B細胞悪性リンパ腫に対しては副作用のより少ない抗がん剤が求められていた。 B-cell malignant lymphoma is a type of non-Hodgkin's malignant lymphoma. Non-Hodgkin's malignant lymphoma includes B-cell malignant lymphoma and T-cell malignant lymphoma, but the incidence of B-cell malignant lymphoma may be overwhelmingly higher than T-cell malignant lymphoma depending on the race and region. The symptom is swollen lymph nodes, but it is generally painless and worsens without subjective symptoms. Since B-cell malignant lymphoma is systemic, it is generally treated with multi-drug chemotherapy, depending on the age and severity of the patient. Therefore, since the burden on patients due to side effects of anticancer drugs is large, anticancer drugs with fewer side effects have been demanded for B-cell malignant lymphoma.
本発明者らは、先ずHMGB1がB細胞悪性リンパ腫の増殖を濃度依存的に高めることを見出した上で、さらに抗HMGB1モノクローナル抗体がB細胞悪性リンパ腫の増殖を有意に抑制することを確認した。なお、T細胞悪性リンパ腫についても同様の実験を行ったが、抗HMGB1抗体によりかえって増殖する場合があった。よって、現段階における本発明者らの知見によれば、抗HMGB1抗体による細胞増殖の抑制効果は、B細胞悪性リンパ腫に特異的なものであると考えられる。 The present inventors first found that HMGB1 increases the proliferation of B cell malignant lymphoma in a concentration-dependent manner, and further confirmed that the anti-HMGB1 monoclonal antibody significantly suppresses the proliferation of B cell malignant lymphoma. A similar experiment was conducted for T-cell malignant lymphoma, but it sometimes proliferated by anti-HMGB1 antibody. Therefore, according to the knowledge of the present inventors at the present stage, it is considered that the effect of suppressing cell proliferation by the anti-HMGB1 antibody is specific to B cell malignant lymphoma.
抗HMGB1モノクローナル抗体は、炎症反応などに関与すると考えられているHMGB1へ選択的に作用し、その作用機序は明らかではないが、B細胞悪性リンパ腫の増殖を抑制する。その一方で、基本的に他のケミカルメディエーター等には作用しない。よって、副作用が生じる可能性はないか、極めて少ないと考えられる。 The anti-HMGB1 monoclonal antibody selectively acts on HMGB1, which is considered to be involved in inflammatory reactions, and the mechanism of action is not clear, but suppresses the growth of B cell malignant lymphoma. On the other hand, it basically does not act on other chemical mediators. Therefore, it is thought that there is no possibility that a side effect will occur or very little.
抗HMGB1モノクローナル抗体の調製は、常法に従えばよい。例えば、市販のHMGB1を用いてマウスやラット等を免疫し、その抗体産生細胞や脾細胞と骨髄腫細胞とを融合させてハイブリドーマを得る。このハイブリドーマをクローニングし、HMGB1へ特異的に反応する抗体を産生しているクローンをスクリーニングする。このクローンを培養し、分泌されるモノクローナル抗体を精製すればよい。 The anti-HMGB1 monoclonal antibody may be prepared according to a conventional method. For example, a mouse, a rat, or the like is immunized using commercially available HMGB1, and the antibody-producing cells, spleen cells, and myeloma cells are fused to obtain hybridomas. The hybridoma is cloned, and a clone producing an antibody that specifically reacts with HMGB1 is screened. This clone may be cultured and the secreted monoclonal antibody may be purified.
本発明で使用する抗HMGB1モノクローナル抗体の種類は、特に制限されない。例えば、ヒト型抗体や完全ヒト抗体を用いることができる。 The type of anti-HMGB1 monoclonal antibody used in the present invention is not particularly limited. For example, a human type antibody or a fully human antibody can be used.
本発明に係るB細胞悪性リンパ腫治療薬の剤形は特に問わないが、有効成分である抗HMGB1モノクローナル抗体がペプチドであることを考慮すれば、注射剤としての投与を志向して、溶液やエマルション製剤などの液状製剤とすることが好ましい。 The dosage form of the therapeutic agent for B-cell malignant lymphoma according to the present invention is not particularly limited, but considering that the anti-HMGB1 monoclonal antibody as an active ingredient is a peptide, it is intended to be administered as an injection and is used as a solution or emulsion. A liquid preparation such as a preparation is preferred.
液状製剤の溶媒としては、pHを調整した生理食塩水やグルコース水溶液など、血漿の等張液を用いることができる。また、抗体を塩類等と共に凍結乾燥した場合には、純水、蒸留水、滅菌水等も使用できる。その濃度も通常の抗体製剤のものとすればよく、一般的には0.1〜1mg/mL程度、点滴用では0.02〜0.2mg/mL程度とすることができる。但し、注射剤の浸透圧は、血漿と同等にする必要がある。 As a solvent for the liquid preparation, an isotonic solution of plasma such as physiological saline adjusted with pH or aqueous glucose solution can be used. In addition, when the antibody is lyophilized with salts or the like, pure water, distilled water, sterilized water, or the like can be used. The concentration may be that of a normal antibody preparation, generally about 0.1 to 1 mg / mL, and about 0.02 to 0.2 mg / mL for infusion. However, the osmotic pressure of the injection must be equivalent to that of plasma.
本発明のB細胞悪性リンパ腫治療薬は、単独で或いは他の抗腫瘍薬や治療手段と併用して使用する。他の抗腫瘍薬と併用する場合には、他の抗腫瘍薬の通常の使用量を抑制することができ、結果としてその副作用を軽減できると考えられる。 The therapeutic agent for B-cell malignant lymphoma of the present invention is used alone or in combination with other antitumor drugs or therapeutic means. When used in combination with other anti-tumor drugs, it is considered that the usual amount of use of other anti-tumor drugs can be suppressed, and as a result, its side effects can be reduced.
本発明に係るB細胞悪性リンパ腫治療薬の投与頻度や投与量は、患者の重篤度や年齢などにより適宜調整すればよい。後述する実施例で示す通り、抗HMGB1モノクローナル抗体の濃度が1μg/mLの培地でも十分なB細胞悪性リンパ腫の増殖抑制効果が観察される。よって、投与すべき製剤の濃度や抗HMGB1モノクローナル抗体の投与量は適宜調整すればよいが、例えば1回当たり0.2〜5mg/(kg体重)投与することができる。 The administration frequency and dose of the therapeutic agent for B-cell malignant lymphoma according to the present invention may be appropriately adjusted depending on the severity and age of the patient. As shown in Examples described later, a sufficient B cell malignant lymphoma growth inhibitory effect is observed even in a medium having an anti-HMGB1 monoclonal antibody concentration of 1 μg / mL. Therefore, the concentration of the preparation to be administered and the dose of the anti-HMGB1 monoclonal antibody may be appropriately adjusted. For example, 0.2 to 5 mg / (kg body weight) can be administered at one time.
以下、実施例を挙げて本発明をより具体的に説明するが、本発明はもとより下記実施例により制限を受けるものではなく、前・後記の趣旨に適合し得る範囲で適当に変更を加えて実施することも可能であり、それらはいずれも本発明の技術的範囲に含まれる。 EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples. However, the present invention is not limited by the following examples, but may be appropriately modified within a range that can meet the purpose described above and below. It is also possible to implement, and they are all included in the technical scope of the present invention.
実施例1 抗HMGB1モノクローナル抗体の調製
(a)ラットの免疫
市販のウシ胸腺由来HMGB1とHMGB2との混合物(和光純薬工業社製、コード番号:080−070741)1mg/mLを2mLガラス製注射筒にとり、別の2mLガラス製注射筒にとった等容量のフロイント完全アジュバンドと連結管を通じて徐々に混和することによって、エマルションとした。セボフルレンにより麻酔したラットの後肢足蹠に、得られたエマルションを0.1mLずつ、計0.2mL注射投与した。2週間後、頚静脈から試験採血し、抗体価の上昇を確認した。次いで、腫大した腸骨リンパ節を前記注射投与から5週間後に無菌的に取り出した。得られた2個のリンパ節から、約6×107個の細胞を回収することができた。
Example 1 Preparation of anti-HMGB1 monoclonal antibody (a) Immunization of rat Commercially available mixture of bovine thymus-derived HMGB1 and HMGB2 (manufactured by Wako Pure Chemical Industries, Ltd., code number: 080-070741) 1 mg / mL in a 2 mL glass syringe The mixture was gradually mixed with an equal volume of Freund's complete adjuvant taken in a separate 2 mL glass syringe through a connecting tube to obtain an emulsion. The resulting emulsion was administered by injection into the hind footpads of rats anesthetized with sevoflurane in an amount of 0.1 mL, for a total of 0.2 mL. Two weeks later, test blood was collected from the jugular vein, and an increase in antibody titer was confirmed. The swollen iliac lymph nodes were then aseptically removed 5 weeks after the injection administration. Approximately 6 × 10 7 cells could be recovered from the two lymph nodes obtained.
(b)細胞融合とクローニング
上記腸骨リンパ節細胞とマウスミエローマSP2/O−Ag14(SP2)細胞を、ポリエチレングリコールを用いて融合させ、得られた融合細胞を96穴マイクロプレートに蒔いた。1週間後、最初のELISAスクリーニングを行ない、陽性ウェルについて、ウェスタンブロットにより二次スクリーニングを行なった。陽性を示すウェル細胞を24穴マイクロプレートに移し、細胞をほぼコンフルエントな状態(約2×105個)に殖やしてから、0.5mLの凍結培地(GIT培地にウシ胎児血清を10%とジメチルスルホキシドを10%添加したもの)を用いて、液体窒素中で凍結保存した。この凍結保存細胞を解凍した後、96穴マイクロプレートでクローニングした。
(B) Cell fusion and cloning The iliac lymph node cells and mouse myeloma SP2 / O-Ag14 (SP2) cells were fused using polyethylene glycol, and the resulting fused cells were seeded in a 96-well microplate. One week later, an initial ELISA screening was performed and positive wells were secondary screened by Western blot. Positive well cells are transferred to a 24-well microplate, and the cells are grown to a nearly confluent state (about 2 × 10 5 cells). Then, 0.5 mL of freezing medium (GIT medium containing 10% fetal bovine serum and dimethyl And 10% sulfoxide added) and stored frozen in liquid nitrogen. The cryopreserved cells were thawed and then cloned in a 96-well microplate.
(c)抗体の精製
回転培養装置(Vivascience社製)により上記陽性細胞を2週間大量培養し、濃度2〜3mg/mLの抗体液を得た。この抗体液をアフィニティゲル(インビトロジェン社製、MEP−HyperCel)と中性pH下で混和し、抗HMGB1抗体をゲルへ特異的に結合させた。特異的にゲルに結合した抗体を、グリシン−塩酸バッファー(pH4)により溶出した。溶出液を限外濾過装置により濃縮した後、セファロースCL6Bゲル濾過カラム(直径2cm×長さ97cm)によって、さらに精製した。
(C) Purification of antibody The positive cells were cultured in a large amount for 2 weeks using a rotary culture apparatus (manufactured by Vivascience) to obtain an antibody solution having a concentration of 2 to 3 mg / mL. This antibody solution was mixed with an affinity gel (MEP-HyperCel, manufactured by Invitrogen) under neutral pH to specifically bind the anti-HMGB1 antibody to the gel. The antibody specifically bound to the gel was eluted with glycine-hydrochloric acid buffer (pH 4). The eluate was concentrated by an ultrafiltration device and further purified by a Sepharose CL6B gel filtration column (
得られたモノクローナル抗体は、HMGB1タンパク質のC末端配列である208EEEDDDDE215(Eはグルタミン酸を示し、Dはアスパラギン酸を示す)をエピトープとして特異的に認識する抗体である。例えば、HMGB1に類似するタンパク質としてHMGB2があるが、HMGB2には211以下のDDDDEの配列が存在しないため、本発明に係るモノクローナル抗体はHMGB2に結合せず、HMGB1のみを特異的に認識して結合することができる。 The resulting monoclonal antibody is an antibody that specifically recognizes 208EEEDDDDE215 (E indicates glutamic acid and D indicates aspartic acid), which is the C-terminal sequence of HMGB1 protein, as an epitope. For example, there is HMGB2 as a protein similar to HMGB1, but since HMGB2 does not have a DDDDE sequence of 211 or less, the monoclonal antibody according to the present invention does not bind to HMGB2 and specifically recognizes and binds only to HMGB1. can do.
実施例2 HMGB1による細胞増殖実験
液体培地(ニッスイ社製、製品名「RPMI 1640」)に、別途調製したヒトHMGB1を1×10-10〜1×10-6g/mLの濃度で溶解した。また、比較のためにHMGB1を添加しない培地も用意した。これら培地(1mL)を直径2cmのシャーレに加え、さらにB細胞悪性リンパ腫株であるFL18とFL218(医仁会武田総合病院の大野仁嗣先生より入手)をそれぞれ約20万個/mLの割合で添加した。なお、FL18はヒトB細胞悪性リンパ腫株であり、FL218は、アポトーシスの抑制作用を有するBcl−2遺伝子が欠損したFL18である。これらB細胞悪性リンパ腫株を37℃で48時間培養した後、各培地における細胞の濃度を顕微鏡で観察することにより測定した。FL18の結果を図1(1)に、FL218の結果を図1(2)に示す。なお、図1中の「*」は、得られた結果をDunnet’sテストにおいて検定した場合にp<0.05で有意差がある場合を示し、「**」はp<0.01で有意差がある場合を示す。
Example 2 Cell Proliferation Experiment with HMGB1 Separately prepared human HMGB1 was dissolved in a liquid medium (manufactured by Nissui, product name “RPMI 1640”) at a concentration of 1 × 10 −10 to 1 × 10 −6 g / mL. Moreover, the culture medium which does not add HMGB1 was also prepared for the comparison. These culture media (1 mL) were added to a petri dish having a diameter of 2 cm, and B18 malignant lymphoma strains FL18 and FL218 (obtained from Prof. Hitoshi Ono of Ichinaikai Takeda General Hospital) were added at a rate of about 200,000 cells / mL, respectively. FL18 is a human B-cell malignant lymphoma strain, and FL218 is FL18 deficient in the Bcl-2 gene having an inhibitory effect on apoptosis. These B cell malignant lymphoma lines were cultured at 37 ° C. for 48 hours, and then the cell concentration in each medium was measured by observing with a microscope. The result of FL18 is shown in FIG. 1 (1), and the result of FL218 is shown in FIG. 1 (2). Note that “*” in FIG. 1 indicates that there is a significant difference at p <0.05 when the obtained result is tested in Dunnet's test, and “**” is p <0.01. The case where there is a significant difference is shown.
図1の通り、HMGB1は濃度依存的にB細胞悪性リンパ腫Bの増殖を促進し、特に1×10-7〜1×10-6g/mLの濃度で増殖速度を有意に高めることが分かった。 As shown in FIG. 1, it was found that HMGB1 promotes the growth of B cell malignant lymphoma B in a concentration-dependent manner, and significantly increases the growth rate particularly at a concentration of 1 × 10 −7 to 1 × 10 −6 g / mL. .
実施例3 抗HMGB1モノクローナル抗体による細胞増殖抑制実験
実施例1で得た抗HMGB1モノクローナル抗体を1μg/mLの濃度で液体培地に溶解した以外は上記実施例2と同様にして、2種のB細胞悪性リンパ腫株を37℃で48時間培養した。培養開始直後、並びに培養開始から2時間後、6時間後、18時間後、24時間後および48時間後における各培地の細胞の濃度を、上記実施例2と同様に測定した。FL18の結果を図2(1)に、FL218の結果を図2(2)に示す。
Example 3 Cell Proliferation Inhibition Experiment with Anti-HMGB1 Monoclonal Antibody Two types of B cells were prepared in the same manner as in Example 2 except that the anti-HMGB1 monoclonal antibody obtained in Example 1 was dissolved in a liquid medium at a concentration of 1 μg / mL. The malignant lymphoma line was cultured at 37 ° C. for 48 hours. The concentration of cells in each medium was measured in the same manner as in Example 2 immediately after the start of culture and at 2 hours, 6 hours, 18 hours, 24 hours and 48 hours after the start of culture. The result of FL18 is shown in FIG. 2 (1), and the result of FL218 is shown in FIG. 2 (2).
図2の通り、抗HMGB1モノクローナル抗体はB細胞悪性リンパ腫の増殖を経時的に抑制している。よって、本発明に係る抗HMGB1モノクローナル抗体は、B細胞悪性リンパ腫の治療効果を有することが実証された。 As shown in FIG. 2, the anti-HMGB1 monoclonal antibody suppresses the growth of B cell malignant lymphoma over time. Therefore, it was demonstrated that the anti-HMGB1 monoclonal antibody according to the present invention has a therapeutic effect on B cell malignant lymphoma.
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