PGN-202

Pictured: An illustration of a woman overlooking a field with pill-shaped sheep/Nicole Bean for BioSpace† A medication called VX-548, developed by Vertex Pharmaceuticals, proved superior to placebo in two small Phase II clinical trials for two types of post-surgical pain, according to a study published in The New England Journal of Medicine in early August. Though the effect was modest, “these trials represent an early foray into an exciting new class of drugs in a difficult field,” Mark Wallace, a physician at UC San Diego’s division of pain medicine who was not involved in the research, wrote in an accompanying editorial. The medication heads now to two Phase III trials with 1,000 participants each, plus an open-label study with 250 participants. The results couldn’t come at a better time, as the opioid epidemic continues to ravage the U.S., with 50,000 people dying each year from overdose, and 10 million abusing the highly addictive medications annually. The U.S. Department of Health and Human Services (HHS) officially declared opioid abuse a public health emergency in 2017, but until there is a safe and effective alternative to opioids, acute and chronic pain present a silent scourge of their own. “There is an immense unmet need for new pain medications, even without the opioid epidemic,” Stephen Waxman, a Yale neuroscientist who penned another editorial accompanying the recent NEJM study, told BioSpace. At least one other firm is pursuing the same target as Vertex, while others are exploring different pain-blocking approaches. “There hasn’t been a new class of pain medicine in more than 20 years,” Vertex spokesperson Heather Nichols told BioSpace in an email. “We believe we could redefine the treatment of pain with the first new mechanism of action in decades.” Meanwhile, other companies are looking to make opioids themselves safer by removing their addictive qualities. Vertex Therapeutic Helps Quell Acute Pain Vertex researchers zeroed in on two distinct, repeatable sources of pain: bunionectomy (a type of foot surgery) and abdominoplasty (tummy tuck), both of which necessitate analgesic medication for acute, post-operative pain. The design allowed for a homogenous patient population, where specific pain was triggered by the same injury. A total of 274 participants were enrolled in the bunionectomy trial, and 303 participants in the abdominoplasty trial. Participants rated their pain at 19 time points on a scale of zero (no pain) to 10 (the worst pain one can imagine). This led to an overall score known as sum of the pain-intensity difference, or SPID, a measurement the FDA recommends as a primary endpoint in acute pain trials. Though a moderate dose of VX-548 did not show a significant improvement over placebo, a high dose did. In fact, the high dose acted rapidly, within the first hour of treatment. At 48 hours, the high dose offered more patients at least a 30% reduction in the score on the pain rating scale than placebo did. Side effects such as nausea, headaches or constipation were mild or moderate, and temporary. Participants on the high dose were less likely to stop medication early because they felt it wasn’t helping than patients on placebo or given a common painkiller that combines hydrodocone bitartrate and acetaminophen. The new molecule is the end result of decades of research into peripheral voltage-gated sodium channels, often referred to as NaV channels, which are found in sensory neurons outside the brain and are responsible for detecting painful or noxious stimuli. There are nine different types of voltage-gated sodium channels in the body (NaV1.1 to NaV1.9); Vertex’s new medication targets NaV1.8, which is expressed exclusively in sensory neurons of the peripheral nervous system and not the brain. Researchers have raised concerns about the results, including the fact that the trials were small, and that a higher (thus more effective) dose of acetaminophen/oxycodone was not tested. Nonetheless, Waxman, whose work on peripheral voltage-gated sodium channels has been widely published, pointed out that the study was a proof-of-concept trial. “It wasn’t necessarily a home run,” he told BioSpace, “but it showed that blocking peripheral gated sodium channels does reduce pain, without central nervous system side effects and without addictive potential.” Waxman added that he is hopeful that these findings will eventually lead to broadly effective treatments for chronic pain as well. Vertex is already conducting a Phase II trial on diabetic neuropathy, with a study completion date in spring of 2024. Other Approaches in Development for Pain Management Vertex is not alone in its quest. “It’s been a race for the holy grail,” commented Waxman. “In 2006, we established very clearly that NaV1.8 is a major controller of the repetitive firing of pain signaling in neurons and if you could block it you would reduce pain.” In Finland, Orion Pharmaceuticals is working on developing a non-opioid pain therapy dubbed JMKX000623 that also blocks the NaV1.8 sodium channel. This experimental therapy is in Phase I trials. And here in the U.S., Virpax Pharmaceuticals is looking at other pain targets and approaches. One of its new analgesics is a liposomal, long-acting, injectable formulation of bupivacaine, a potent local anesthetic similar to lidocaine. It is intended for post-operative pain management and Virpax is developing the formulation in conjunction with the U.S. Army, which is studying its potential in battlefield wounds. The company is also planning Phase I safety trials on two intranasal sprays that would target pain centers in the brain—one is a peptide called enkephalin, and the other, a pharamaceutical-grade cannabidiol. Meanwhile, in an entirely different approach, biotech startup Ensysce Biosciences has developed a novel chemistry that modifies how opioids are released when taken in order to make them safer. The platform, called TAAP (trypsin activated abuse protection), uses the body’s own digestive enzyme, trypsin, to trigger release of the opioid. That means the pills won’t work if they crushed in order to be snorted or injected—the kind of recreational abuse that is common. The opioid can only be activated in the stomach in the presence of trypsin. In addition, overdose protection is ensured by adding a tiny amount of trypsin inhibitor to every dose. “If you take the prescribed amount, the amount of trypsin inhibitor is negligible so nothing happens,” explained chemist Lynn Kirkpatrick, CEO at Ensysce Biosciences. “But if you take more than you should, there is enough inhibitor to shut down the activation.” The company’s molecule is called PF614 and is currently in Phase II clinical trials, and fast tracked by the FDA. It is a prodrug of oxycodone, with a safety pro to other traditional opioids, but a 12-hour half-life once ingested, which is longer-acting than current opioids. “We are talking to the FDA about Phase III plans,” Kirkpatrick told BioSpace. If all goes as planned, she said, the company might have an NDA submitted to the agency by 2025. Jill Neimark is a freelance science writer based in Macon, Georgia. Reach her at jillneimark.com.

肿瘤免疫是一个不断发展的研究领域，涉及肿瘤微环境中的免疫细胞。中性粒细胞胞外陷阱(NETs)是中性粒细胞衍生的胞外网状染色质结构，由组蛋白和颗粒蛋白组成。NET最初被发现是宿主对病原体的主要防御，由于它与肿瘤密切相关，越来越受到人们的关注。NET的过度形成与肿瘤生长、转移和耐药性的增加有关。此外，通过对免疫细胞的直接和/或间接影响，NETs的异常增加有利于免疫排斥和抑制T细胞介导的抗肿瘤免疫应答。近期的一项研究，首次报道了NETosis在肿瘤耐药方面的作用及其具体机制。2023.4.10在Cancer Cell IF 50.3/Q1发表了一篇文章，题为“在化疗过程中形成的中性粒细胞细胞外陷阱通过TGF-b激活赋予治疗耐药性 Neutrophil extracellular traps formed during chemotherapy confer treatment resistance via TGFb activation”作者发现了一种“化疗NETosis通路”，其中化疗诱导的NETosis是恶性肿瘤中产生化疗耐药的关键角色。为了探讨化疗如何重塑肺转移的炎症微环境，作者首先将410.4小鼠乳腺癌细胞静脉注射Balb/c小鼠，获得肺转移模型。在接受顺铂或阿霉素/环磷酰胺(A/C)化疗后，他们评估了肺转移模型中的中性粒细胞浸润水平，并观察到化疗后中性粒细胞浸润水平升高。此外，他们发现中性粒细胞耗竭改善了化疗反应。该项研究表明，化疗促进中性粒细胞细胞外陷阱(NET)的形成，降低了治疗乳腺癌肺转移的化疗疗效。NETs在癌细胞中诱导TGF-b依赖的上皮-间充质转化(EMT)， NET/ TGF-b轴的药理学靶向改善了化疗疗效。阐述了化疗诱导的炎症如何通过促进恶性肿瘤中的NETosis而赋予化疗耐药，强调了在癌症治疗中靶向炎性NETs的治疗机会。一背景中性粒细胞细胞外陷阱(NETs)由含有各种蛋白酶和整合素(ITGs)的NET-DNA支架组成，在各种压力下从中性粒细胞中释放出来，这一过程被称为NETosis。最近，已经证明NETs可以通过细胞表面DNA传感器CCDC25直接吸引肿瘤细胞，隔离循环肿瘤细胞或唤醒转移前生态位中的休眠肿瘤细胞，从而促进肿瘤转移虽然恶性肿瘤中NET的形成与放射治疗耐药之间的联系已被报道，但NETosis是否可能由化疗诱导并有助于化疗应答尚不清楚。二主要结果作者提出了一种机制，在化疗反应中，中性粒细胞浸润肿瘤微环境并形成NETs，导致肿瘤化疗耐药。化疗在肺转移灶中上调CXCL1和CXCL5，导致中性粒细胞募集。在细胞毒性药物的作用下，垂死的肿瘤细胞释放出大量的ATP，邻近肿瘤细胞上的嘌呤能受体P2RX7感知到ATP，并激活NLRP3炎症小体和caspase-1将pro-IL-1b裂解为IL-1b。由IL-1b触发，浸润的中性粒细胞在肺转移中发生NETosis。随后，NET成分ITGavb1捕获肿瘤细胞不断产生的潜伏TGF-b，并通过基质金属蛋白酶9 (MMP9)促进其激活。最后，活性TGF-b与肿瘤细胞上的TGF-bR1结合，引起上皮-间充质转化(EMT)和化疗耐药。此外,顺铂治疗后，癌细胞中CXCL5和CXCL5上调，CXCL1/5-CXCR2轴参与了化疗后将中性粒细胞招募到转移性肺中为了研究NET对化疗敏感性的贡献，作者用含有NET的条件培养基(CM)治疗乳腺癌细胞，使肿瘤细胞具有化疗耐药性。通过PAD4抑制剂或DNase I去除NETs显著改善化疗反应，表明NETosis和化疗耐药之间存在直接因果关系。在接受化疗的转移性乳腺癌患者血液样本中，血浆NET水平与转移的化疗耐药相关，提示血浆NET可作为乳腺癌化疗患者预后的生物标志物。为了研究这些观察的机制基础，作者对化疗后的癌细胞CM进行了蛋白质组细胞因子检测，结果证实了IL-1b的上调。虽然IL-1b在触发NETosis中的作用已有报道，但IL-1b在触发NETosis中的关键作用在耐药方面的报道尚属首次。IL-1b阻断后，顺铂治疗小鼠肺转移和肾脏的NETosis均减少，化疗反应和肾功能均有显著改善。有趣的是，IL-1b不是由中性粒细胞分泌的，而是由邻近的肿瘤细胞分泌的。强调了肿瘤微环境中免疫细胞、基质细胞和癌细胞之间的相互作用。邻近的肿瘤细胞通过嘌呤能受体P2X7 (P2RX7)对死亡肿瘤细胞释放的“危险信号”ATP做出反应，从而触发NLRP3炎症小体和caspase-1的级联激活，最终将pro-IL-1b裂解为IL-1b。在随后对NETs CM处理的410.4乳腺癌细胞的RNA测序分析中，发现TGF-b和上皮-间充质转化(EMT)相关信号通路上调。在肺组织中，只有在NETs存在的情况下，才会在癌细胞中观察到EMT特征的增加，而靶向NETs或TGF-b受体1 (TGF-bR1)则会取消这种转变。ELISA检测发现，410.4肿瘤细胞的CM中存在大量laTenTTGF-b，而中性粒细胞或含NETs的CM中则不存在。有趣的是，当混合前一种和后一种CMs时，active TGF-b的数量急剧增加，而抵消NETs则显著地取消了潜在的TGF-b激活。此外，作者揭示了NETs通过降解潜伏相关蛋白(LAP)释放活性TGF-b, LAP是潜伏- TGF-b的小潜伏复合体(SLC)形式的一部分。此外，通过使用针对主要NETs相关蛋白酶的不同抑制剂——中性粒细胞弹性蛋白酶(NE)、基质金属蛋白酶9 (MMP9)、组织蛋白酶G和髓过氧化物酶(MPO)——作者发现，只有MMP9抑制剂可以消除NET诱导的TGF-b激活、化疗耐药和EMT。总之，这些发现揭示了NET相关的MMP9负责激活肿瘤细胞分泌的潜伏TGF-b，随后导致肿瘤细胞EMT和化疗耐药。为了研究靶向IL-1b/TGF-b轴的临床转化，作者将注意力转向了ITGavb1的功能，因为ITGavb1可以通过延迟相关肽-精氨酸-甘氨酸-天冬氨酸(LAP-RGD)结构域结合潜伏- TGF-b。他们发现ITGavb1对于激活潜伏的TGF-b至关重要，并且抵消ITGavb1可以改善化疗反应。令人惊讶的是，免疫荧光结果显示TGF-b被捕获在NET-DNA支架内，而阻断ITGavb1则阻止了捕获。NETs在化疗相关并发症的作用。急性肾损伤(AKI)是化疗的严重并发症，可导致显著的发病率和死亡率。NETs的形成已被揭示通过诱导小管坏死和损伤组织来加剧AKI，其中NET相关组蛋白被报道以TLR2/4依赖的方式引起小管损伤在顺铂诱导肾毒性的背景下，作者表明，在顺铂治疗后，NET在肾脏中形成，与肿瘤负荷无关，这表明靶向NETs除了有利于化疗疗效外，还有助于缓解化疗肾并发症。作者具有启发性的发现强调了化疗诱导炎症的“阴暗面”，并提出了一些需要进一步研究的重要问题。首先，DAMP(包括ATP)信号诱导的炎症是一把双刃剑;除了引起EMT和肿瘤细胞的化疗耐药外，它还可以通过作用于其他免疫细胞(即DC)来增强免疫应答。因此，需要更全面和深入地认识化疗诱导的炎性微环境中各种DAMPs的双重性。原发肿瘤组织中肿瘤分泌的CXCL1和CXCL5较少，而转移性肺组织中肿瘤分泌的CXCL1和CXCL5较多的原因仍有待阐明。此外，NETs/ITGavb1/MMP9/TGF-b轴的激活是否也在介导其他癌症类型的化疗耐药中发挥作用仍有待研究。更重要的是，这项有趣的工作揭示了除了受损肿瘤细胞释放的DAMPs外，NETs也应被视为肿瘤中的危险信号。更好地了解炎症的后果和肿瘤微环境中的各种危险信号将有助于合理设计抗癌疗法。三总结和思维扩展这篇文章的高光部分在于：化疗促进乳腺癌肺转移中NET的形成；NETs促进乳腺癌肺转移化疗耐药性；NETs激活潜伏的TGF-b，促进癌细胞中的EMT；NETs在化疗相关并发症的发挥作用；靶向NETs改善化疗对肺转移的疗效；发现了吗，研究肿瘤化疗耐药机制的思路又又又更新了，以上五处高光思路，处处有新意，可以通过整合公共数据库细胞系、肿瘤队列等数据，自由创作：1. 筛选化疗耐药gene signature ,预后模型、耐药预测模型构建；2.化疗相关并发症诊断和预测；3.EMT在化疗耐药中发挥的作用、异质性、及其与NETs之间的互作机制；4.改善肿瘤化疗敏感性的靶标鉴定和疗效预测；总之，化疗耐药，一直是临床上高居不下的热点之一，但是耐药机制分析往往需要做大量的基础实验，科研小白无论是精力还是财力，真的是“鸭梨山大”，能够运用纯生信来探索耐药机制的思路，真是寥寥无几，凤毛麟角，错过这一波，恐怕下一波有遥遥无期啦！看见的小伙伴，个性化的耐药机制分析等着你，赶快上车吧！封面图来源：网络倒计时三周丨PHDI 2023 制药产业数智化峰会最新议程&大咖阵容公布诺和诺德中国首届研发日——畅想没有严重慢性疾病的未来【9月6日，药时代直播间，不见不散！】点击这里，与药时代一起快乐学习！