Apparatus and method for priming solid tumors with pressure pulses to enhance anti-cancer therapy

Abstract

The present disclosure relates to devices and methods for priming the tumor microenvironment with pressure pulses to enhance the efficacy of anti-cancer therapeutics in subjects in need thereof. Additionally, the disclosure also discloses an increased response of solid tumors exposed locally to stress waves to systemically administered therapeutics. An apparatus according to the present disclosure includes: (1) a pulsed laser system; (2) an optical guide that directs laser pulses to one or more optical pressure transducers; (3) one or more optical pressure transducers that absorb laser pulses from the pulsed laser system to generate pressure pulses; (4) the one or more optical pressure transducers; (5) a tumor positioning structure configured to engage a solid tumor; and (6) a control system that limits exposure of the solid tumor to the pressure pulses. The anti-cancer therapeutic agent can be administered before, after, or during pressure pulse priming of the solid tumor.

Description

[Technical Field]

[0001] The present disclosure relates generally to the field of tumor microenvironment (TME) remodeling (or priming) to enhance the efficacy of anti-cancer therapeutics. More specifically, the present disclosure relates to a pressure pulse tumor priming device and methods of using the same, as well as methods of using pressure pulses to prime the tumor microenvironment, thereby enhancing the efficacy of anti-cancer therapeutics. [Background technology]

[0002] Active stimulation of the host immune system to induce antitumor immune responses capable of controlling tumor growth and metastasis has been pursued for a century and has recently reached the status of first-line cancer treatment [1-3]. A notable example is immune checkpoint blockade (ICB) therapy, which uses monoclonal antibodies (mAbs) to block inhibitory immune receptors (checkpoints) and unlock the antitumor functions of immune cells present in the immunosuppressive tumor microenvironment (TME). To promote a therapeutic response, CD8+ T cells, strongly activated by tumor antigens, must be prevented from being suppressed by negative regulators. These negative regulators are called "checkpoints" because they detect, resist, and reverse overreactions. While checkpoint inhibitors are essential for maintaining self-tolerance and preventing autoimmune disease, they can also be hijacked by tumor cells to escape immune surveillance. Inhibitory immunoreceptors include, but are not limited to, CTLA4 (cytotoxic T lymphocyte-associated protein 4), PD1 (programmed cell death protein 1), LAG3 (lymphocyte-activation gene-3), TIM3 (T-cell immunoglobulin and mucin domain-3), TIGIT (T-cell immunoreceptor with Ig and ITIM domains), ICOS (inducible T-cell costimulatory receptor), BTLA (B- and T-lymphocyte attenuator), and VISTA (V-domain Ig-containing suppressor of T-cell activation). CTLA4 and PD1 are the most potent examples of T-cell immune checkpoint molecules currently known.

[0003] The clinical approval history of ICBs by the U.S. Food and Drug Administration (FDA) is impressive [4]: ​​ipilimumab (2011), pembrolizumab (2014), nivolumab (2014), atezolizumab (2016), durvalumab (2017), avelumab (2017), cemiplimab (2018), dostarlimab (2021), tislelizumab (2021), and leratilimab (2022). Other ICBs, such as toripalimab (2018), camrelizumab (2019), and sintilimab (2019), have also been approved for clinical use in China. Furthermore, some of these ICBs have also been approved for the treatment of different types of cancer. Due to this significant advancement in immunotherapy, it was estimated that approximately 44% of cancer patients in the United States were eligible for treatment with ICB in 2019. However, more than 87% of cancer patients do not respond to ICB.[5] There is a critical need to improve the efficacy of immunotherapy.

[0004] CTLA4 molecules are contained in intracellular vesicles of naive T cells and are constitutively expressed on the surface of CD4+CD25+ regulatory T (Treg) cells. Naive T cells are activated when their T cell receptors bind specific antigens presented by antigen-presenting cells (APCs) in the presence of costimulatory signals. This costimulatory signal is the binding of CD28 expressed on the T cell surface to B7 molecules (B7.1, also known as CD80, or B7.2, also known as CD86) on APCs. APCs are immune cells that process antigens and present them for recognition by T cells, including B lymphocytes, dendritic cells, macrophages, and other immune cells. CD28 and CTLA4 compete for binding to B7-1 and B7-2 on APCs, but CTLA4 binds more strongly to B7-1 and B7-2, transmitting a negative signal to T cells rather than a costimulatory signal. CTLA4 antagonizes multiple internal signaling pathways, preventing T cell activation and proliferation [6]. Treg cells, which constitutively express CTLA4, can silence T cell responses. The recognition that CTLA4 is a negative regulator of T cell activity suggested that blocking its action could restore T cell responses to cancer cells. Indeed, neutralizing anti-CTLA4 mAbs enhance antitumor immunity. In addition to promoting CD8+ effector T cell responses, anti-CTLA4 therapy also reduces local Treg cells within tumors through antibody-dependent cell-mediated cytotoxicity, shifting the balance of the TME away from immunosuppression [7].

[0005] Human PD1 is expressed on T cells after T cell receptor stimulation and binds to PDL1 and PDL2, B7 homologs that are constitutively expressed on APCs and can be induced in nonhematopoietic tissues. PDL1 (also known as programmed cell death ligand 1, B7-H1) is a transmembrane protein that downregulates immune responses through binding to its two inhibitory receptors, PD1 and B7.1, and is present much more frequently than PDL2 (also known as programmed cell death ligand 2, B7-DC), on many cell types, including T cells, tumor cells, epithelial cells, and endothelial cells. PD1 suppresses immune responses primarily through inhibitory signaling in CD8+ effector T cells and Treg cells [7]. Binding of PD1 to its ligand can induce a state of T cell dysfunction known as T cell exhaustion. Tumor cells can upregulate PD1 ligands. PDL1 expressed by cells within the TME binds to PD1 on T cells, subsequently triggering inhibitory signaling, blocking effector function and reducing the killing ability of T cells. Therefore, tumor cells can induce T cell exhaustion and generate a TME that promotes tumor growth and invasion. Because the PD1 / PDL1 pathway protects cells from T cell attack, anti-PD1 and anti-PDL1 antibodies can improve the functional properties of CD8+ effector T cells at tumor sites.

[0006] T cell activation allows T lymphocytes to recognize antigens on specific target cells. Activated CD8+ T cells gradually transform into effector T cells (or cytotoxic T lymphocytes, CTLs), which recognize and kill target cells via different pathways. In one pathway, Fas ligand (or CD95L) expressed on the surface of CTLs binds to Fas receptor (or CD95) on the target cell, inducing apoptosis via the caspase cascade. In another pathway, CTLs release granulysin, perforin, cathepsin C, and / or granzymes into the intercellular space between the CTL and the target cell, which are highly cytotoxic to the target cell. Although the mechanisms of these immunotherapies are complex and interrelated and are still under investigation, these pathways indicate that CTL infiltration into solid tumors is necessary for antitumor activity. The same requirement for infiltration into the TME also applies to other immunotherapies, such as adoptive T cell therapy and cancer vaccines.

[0007] The failure of immunotherapy in the majority of patients, and its remarkable success in a subset, has been the subject of intense scrutiny. The lack of therapeutic efficacy has been attributed to a variety of factors, including an abnormal tumor mesothelial cell membrane (TME), characterized by dysfunctional blood vessels that impede the delivery of immunotherapeutic drugs and cause immunosuppression. Indeed, the spatiotemporal lack of adequate tumor perfusion can result in hypoxia, low pH, and inadequate drug delivery, potentially compromising the efficacy of cancer treatments, including immunotherapy. The difficulty of anticancer drugs accessing all cells within the TME has been recognized in chemotherapy [8] and is expected to become even more limiting for biologics and immunotherapy, given the size of the corresponding therapeutics. Rapid cancer cell proliferation distends blood vessels within solid tumors, reducing vascular density, compressing blood and lymphatic vessels, limiting oxygen and nutrient supply, accumulating metabolic products, and lowering pH. The difficulty of infiltrating solid tumors is exacerbated when mAbs are used because they are very large macromolecules, often aggregates of macromolecules with molecular weights of approximately 150 kDa. The physicochemical properties and large size of mAbs preclude passive diffusion through vascular epithelial cells. The primary mechanism by which mAbs are distributed throughout the body and into tissues is convective transport, i.e., via the blood-tissue hydrostatic pressure gradient. The affinity of mAbs for target antigens in the interstitial space or on cell surfaces and their convection into lymphatics determine their retention in tissues. The efficiency of convection into the interstitial space is much lower than that from the interstitial space. As a result, the clinical volume of the central compartment of most mAbs ranges from 2 to 3 L, similar to vascular water, with an overall volume of distribution at steady state of 8 to 20 L [9]. The limited volume of distribution of mAbs, especially ICBs, indicates that they are primarily confined to the vascular compartment

[10] .

[0008] Increased vascular leakage in solid tumors and the inability of intratumoral lymphatics to effectively drain this fluid have been explored by nanoparticle delivery systems, exploiting the enhanced permeability and retention effect to accumulate therapeutic agents within tumors.

[11] Nanoparticle drug carriers have been shown to mediate greater tumor drug deposition compared to free drugs, and the delivered drug can persist for several days at concentrations exceeding the maximum tumor concentration achieved with free drugs. However, nanomedicines tend to accumulate in reticuloendothelial tissues (spleen, liver, and lungs) and remain near tumor vascular openings.

[12] Nanomedicines have a low probability of reaching the majority of target cells within tumors.

[0009] The mechanical microenvironment of solid tumors is characterized by elevated interstitial fluid pressure (IFP) and solid stress. These two stressors in the TME are distinct in that IFP is an isotropic stress from fluid due to increased leakage and poor drainage within the tumor, whereas solid stress is exerted by non-fluid components. IFP results from the high vascular permeability of the TME and mechanical compression of downstream blood and lymphatic vessels. Solid stress is associated with the hyperproliferation of cancer cells, which exert forces on structural elements in neighboring tumor and normal tissues, resulting in equal and opposite forces according to the law of action and reaction. Vascular compression has two effects on therapeutic agents: (i) vascular collapse hinders drug and immune cell access, and (ii) lack of lymphatic function reduces drainage and increases IFP, reducing the transport of large therapeutic agents (e.g., antibodies that block inhibitory checkpoint molecules) and nanomedicines, which are diffusive and therefore less effective due to their large size. These effects are further exacerbated in immunotherapies that rely on the infiltration of tumor antigen-specific cytotoxic T lymphocytes into solid tumors, including adoptive T cell therapy and cancer vaccines.

[0010] Abnormal and disorganized tumor vasculature is a target for various therapeutic approaches, including antiangiogenic agents such as anti-vascular endothelial growth factor (anti-VEGF). At low doses, antiangiogenic treatments normalize tumor vasculature, improving tumor perfusion and drug delivery

[13] . Treatments aimed at overcoming the IFP and high growth-induced solid stress of solid tumors are less common. In normal tissue, IFP ranges from 0 to 3 mmHg. However, due to fluid leakage from blood vessels in solid tumors and the inability of intratumoral lymphatics to effectively drain this fluid, IFP can range from 5 to 40 mmHg, even reaching 75 to 130 mmHg in dermal-forming pancreatic tumors

[14] , with a rapid drop at the tumor margin. Anti-solid stress strategies differ from vascular normalization strategies, which use antiangiogenic agents to organize immature vessels and strengthen remaining vessels. Anti-solid stress strategies aim to decompress blood vessels and increase perfusion

[15] . If venous resistance is reduced and lymphatic drainage is reestablished, IFP also decreases. IFPs and solid stress limit the ability of anticancer drugs to reach tumor cells within solid tumors. The delivery of immune cells to solid tumors is even more severely restricted. This can dramatically impact the efficacy of immunotherapy. In particular, CD8+ T cells can be excluded from or trapped within tumors by the dense fibrous extracellular matrix produced by cancer-associated fibroblasts

[16] . CTL infiltration into tumors correlates with the therapeutic efficacy of ICB.

[0011] As a pharmacological approach to alleviating solid tumor stress, tumor-targeted angiotensin receptor blockers have been shown to reduce the activity of cancer-associated fibroblasts and showed enhanced efficacy when combined with ICBs.

[17] For example, the combination of angiotensin receptor blockers with an ICB mixture of aCTLA-4 and aPD-1 extended the median survival time of mice bearing orthotopic 4T1 tumors from 17 days in the control group to 24 days in the combination group (median survival time for mice treated with aCTLA-4 and aPD-1 was 20 days)

[17] . While such pharmacological approaches can reduce solid tumor stress and enhance the efficacy of immunotherapy, they also have systemic effects.

[0012] One might think that the effects of high IFP and solid stress are less important for chemotherapy using small-molecule drugs because of their high diffusion coefficients. However, this consideration ignores the fact that most small-molecule drugs have low or no solubility in aqueous media and bind extensively to plasma proteins such as albumin and low- and high-density lipoproteins immediately after administration. For example, human serum albumin (HSA) has a molecular weight of approximately 67 kDa, nearly half the molecular weight of a monoclonal antibody. However, small-molecule drugs bound to HSA may have difficulty penetrating solid tumors, similar to macromolecules, nanomedicines, biologics, or CTLs.

[0013] Considering that the effects of vascular normalization and solid stress reduction are interrelated and both contribute to improved drug delivery to solid tumors, the term "tumor priming" has been proposed to refer to these strategies

[14] . Drug penetration into solid tumors has also been enhanced by photodynamic tumor priming, which uses a photosensitizer at subtumor-killing concentrations to improve tumor permeability to chemotherapeutic and biologic drugs

[18] . While this allows for spatiotemporal control of solid tumor permeability with low-toxicity photosensitizers, it remains a pharmacological approach requiring the use of additional pharmaceutical agents. Acoustic priming of solid tumors using focused ultrasound has also been reported

[19] . The application of focused ultrasound to biological tissues has been associated with the generation of thermal and cavitational effects that alter the physiology of target cells

[19] .

[0014] Acoustic priming therapy is performed at a frequency of 10 to 1000 W / cm2 in the treatment area. 2 Using a piezoelectric transducer configured to generate ultrasound waves at frequencies between 0.01 and 10 MHz with a spatial peak temporal average intensity (ISPTA) of 100 W / cm, ultrasound is applied continuously to a specific volume of the treatment area for a duration ranging from 0.5 to 5 seconds.

[19] The ISPTA value for acoustic priming therapy is within the current FDA power limit for diagnostic ultrasound: ISPTA < 0.72 W / cm. 2 is significantly exceeded.

[0015] Current pharmacological approaches to solid tumor priming have met with some success but are not without off-target effects. Radiological approaches to solid tumor priming increase the body's exposure to harmful radiation. Acoustic tumor priming requires ultrasound waves with peak-to-peak, time-averaged spatial intensities significantly exceeding the FDA-established power safety limits for diagnostic ultrasound. This disclosure describes, for the first time, the use of pressure pulses with peak-to-peak, time-averaged spatial intensities within the FDA-established power safety limits for diagnostic ultrasound to achieve solid tumor priming and increase tumor response to therapeutic agents. Summary of the Invention [Means for solving the problem]

[0016] The use of pressure pulses in tumor priming therapy (PPTPT) is disclosed herein for the first time. In one embodiment, PPTPT uses laser light pulses and piezoelectric optical (light pressure) transducers to generate high-pressure broadband photoacoustic waves that cross solid tumors and enhance the penetration of chemotherapeutic agents, biologics, immunologics, or any combination thereof. In another embodiment, PPTPT uses laser light pulses to ablate the surface of material, generating shock waves that pass through solid tumors and enhance the penetration of chemotherapeutic agents, biologics, immunologics, or any combination thereof within the tumor. PPTPT combines exposing solid tumors to pressure pulses with the administration of chemotherapy, biologics, or immunotherapy. The pressure pulses act primarily by priming solid tumors, allowing for deeper penetration of chemotherapeutic agents, biologics, or immunologics into the solid tumor, resulting in a more robust tumor response to the chemotherapeutic agents, biologics, immunologics, or any combination thereof.

[0017] In one aspect of the present disclosure, an apparatus for tumor priming with pressure pulses is provided, comprising: a pulsed laser system having a pulse repetition rate of 0.1 Hz to 100 Hz; an optical guide configured to direct laser pulses to one or more optical pressure transducers; one or more optical pressure transducers configured to absorb laser pulses from the pulsed laser system and generate pressure pulses, wherein the pressure pulses have a peak compressive pressure of 0.1 MPa to 100 MPa and 90% of each pressure pulse lasts for 0.1 ns to 500 ns; a tumor positioning support structure configured to couple the one or more optical pressure transducers to a selected region of a solid tumor at a distance of less than 3 cm from the region; and a control system configured to limit exposure of the solid tumor to the pressure pulses for a time period of 1 second to 60 minutes.

[0018] In further embodiments, the light guide comprises one or more optical fibers or light pipes.

[0019] In further embodiments, the light guide comprises a mirror, a lens, a prism, a diffuser, a polarizer, or any combination thereof.

[0020] In a further embodiment, the optical pressure transducer comprises a laser light absorption system and a material with a Gruneisen parameter higher than 0.5, and each pressure pulse is a photoacoustic wavefront.

[0021] In a further embodiment, the optical pressure transducer is a laser light absorption system and a 200 mJ / cm 2 With a material having an ablation threshold below, each pressure pulse is a shock wave front.

[0022] In a further embodiment, the tumor localization support structure is configured to hold one or more optical pressure transducers with an acoustic coupling element disposed between the transducers and the surface of the solid tumor.

[0023] In a further embodiment, the tumor localization support structure is an endoscope and the light guide is one or more optical fibers configured to carry laser light from a light source through the endoscope to one or more optical pressure transducers at the distal end of the optical fiber.

[0024] In a further embodiment, the endoscope is configured for insertion into a hollow organ through a natural body orifice or a body incision less than 2 cm in length.

[0025] In a further embodiment, the tumor localization support structure is a catheter and the light guide is one or more optical fibers configured to carry laser light from the light source through the catheter to one or more optical pressure transducers at the distal end of the optical fiber.

[0026] In further embodiments, the catheter is configured for insertion into a body cavity, duct, blood vessel, brain, skin, or fatty tissue.

[0027] In a further embodiment, the tumor localization support structure comprises a sharpened tip configured to allow insertion of one or more optical pressure transducers into a solid tumor.

[0028] In another aspect of the present disclosure, a method is provided for treating a solid tumor in a subject afflicted with cancer, the method comprising: pressure-pulse priming the solid tumor by exposing the solid tumor to one or more pressure pulses, wherein the pressure pulses have a peak compressive pressure of 0.1 MPa to 100 MPa and 90% of each pressure pulse lasts for 1 ns to 500 ns; and administering to the subject one or more anti-cancer therapeutic agents, thereby treating the solid tumor in the subject afflicted with cancer. In a further embodiment, the pressure-pulse priming of the solid tumor is performed by an apparatus according to the present disclosure.

[0029] In a further embodiment, the method further comprises repeating the pressure pulse tumor priming, the administration of one or more anti-cancer therapeutic agents, or both, at least once at doses that improve the response of the solid tumor to the treatment.

[0030] In further embodiments, the anti-cancer therapeutic agent is selected from the group consisting of an inhibitor of an inhibitory checkpoint molecule, an activator of a stimulatory checkpoint molecule, an antibody, a cytokine, an interferon, an interleukin, a vaccine, an oncolytic virus, a chimeric antigen receptor T cell, and any combination thereof.

[0031] In a further embodiment, the therapeutic agent is a biotherapeutic agent.

[0032] In a further embodiment, the therapeutic agent is a monoclonal antibody (mAb) used in cancer therapy, or any combination of mAbs used in cancer therapy.

[0033] In further embodiments, the therapeutic agent is selected from ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, cemiplimab, dostallimab, tislelizumab, leratilimab, toripalimab, camrelizumab, sintilimab, or any combination thereof.

[0034] In a further embodiment, the therapeutic agent is a cytostatic or cytotoxic drug bound to a plasma protein.

[0035] In a further embodiment, the therapeutic agent is a macromolecule.

[0036] In a further embodiment, the therapeutic agent is a nanomedicine.

[0037] In another aspect of the present disclosure, there is provided an apparatus for treating a solid tumor in a subject suffering from cancer, the apparatus comprising means for generating pressure pulses having a peak compressive pressure of 0.1 MPa to 100 MPa, with 90% of each pressure pulse lasting between 1 ns and 500 ns.

[0038] In a further embodiment, the device is a device according to the present disclosure.

[0039] In another aspect of the present disclosure, there is provided a system comprising at least one device according to the present disclosure.

[0040] In another aspect of the present disclosure, a kit is provided that includes a device according to the present disclosure and an anti-cancer therapeutic agent.

[0041] In further embodiments, the anti-cancer therapeutic agent is selected from the group consisting of an inhibitor of an inhibitory checkpoint molecule, an activator of a stimulatory checkpoint molecule, an antibody, a cytokine, an interferon, an interleukin, a vaccine, an oncolytic virus, a chimeric antigen receptor T cell, and any combination thereof.

[0042] In a further embodiment, the therapeutic agent is a biotherapeutic agent.

[0043] In a further embodiment, the therapeutic agent is a monoclonal antibody (mAb) used to treat cancer, or any combination of mAbs used to treat cancer.

[0044] In further embodiments, the therapeutic agent is selected from ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, cemiplimab, dostallimab, tislelizumab, leratolimab, toripalimab, camrelizumab, sintilimab, or any combination thereof.

[0045] In a further embodiment, the therapeutic agent is a cytostatic or cytotoxic drug bound to a plasma protein.

[0046] In a further embodiment, the therapeutic agent is a macromolecule.

[0047] In a further embodiment, the therapeutic agent is a nanomedicine.

[0048] Unless otherwise defined, all technical and / or scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this disclosure belongs.In carrying out or testing embodiments of the present disclosure, methods and materials similar or equivalent to those described herein can be used, but exemplary methods and / or materials are described below.In the event of any discrepancy, the patent specification, including definitions, shall prevail.In addition, the materials, methods, and examples are merely illustrative and are not necessarily intended to be limiting.

[0049] Further embodiments and the full scope of applicability of the present disclosure will become apparent from the detailed description set forth below. It should be understood, however, that the detailed description and specific examples, while indicating preferred embodiments of the present disclosure, are given by way of illustration only, and that various changes and modifications within the spirit and scope of the present disclosure will become apparent to those skilled in the art from this detailed description. [Brief explanation of the drawings]

[0050] Several embodiments of the present disclosure are described herein, by way of example only, with reference to the accompanying drawings. Specific reference will be made to the details of the drawings, where it is emphasized that the details shown are exemplary and are for the purpose of explanatory discussion of the embodiments of the present disclosure. In this regard, the description accompanying the drawings will make apparent to those skilled in the art how the embodiments of the present disclosure may be practiced. Dimensions of components and features shown in the figures have been chosen for convenience and clarity and are not necessarily drawn to scale. [Figure 1] Figures 1A-B show absolute pressure pulses generated by a photovoltaic material made of carbon nanoparticles and PDMS when excited with a laser fluence of (Figure 1A) approximately 60 mJ / cm and (Figure 1B) approximately 126 mJ / cm for an 8 ns duration, as detected by a hydrophone calibrated over the 1 to 30 MHz range. [Figure 2]Figure 2 shows the Fourier transform of the stress wave generated by a photovoltaic material made of carbon nanoparticles and polymer when excited with a laser fluence of approximately 60 mJ / cm2 and duration of 8 ns, as measured with a 225 MHz contact transducer. [Figure 3] Figure 3 shows the in vitro viability of immortalized monkey fibroblasts (COS-7) in control (CTR) cell culture plates and plates exposed to photoacoustic waves for 5 minutes (5 min) or 10 minutes (10 min) at laser repetition rates of 6 Hz or 20 Hz. Cell viability was not impaired by exposure to up to 12,000 pressure pulses. [Figure 4] Figure 4 shows magnetic resonance imaging (MRI) of tissue in the neck region of a Sprague-Dawley rat. The left side was exposed to photoacoustic waves for 5 minutes, 5 times per week for 4 weeks, at a pulse repetition rate of 20 Hz, with a peak compressive pressure of approximately 3 MPa. No differences were observed on the left side compared to the right side, which was not exposed to photoacoustic waves. No adverse effects were observed in the area where photoacoustic waves were applied. Figure 4 shows magnetic resonance imaging (MRI) of tissue in the neck region of a Sprague-Dawley rat. The left side was exposed to photoacoustic waves for 5 minutes, 5 times per week for 4 weeks, at a pulse repetition rate of 20 Hz, with a peak compressive pressure of approximately 3 MPa. No differences were observed on the left side compared to the right side, which was not exposed to photoacoustic waves. No adverse effects were observed in the area where photoacoustic waves were applied. [Figure 5] Figure 5 shows hematoxylin and eosin staining of tissue from the left carotid artery region of a Sprague-Dawley rat exposed to photoacoustic waves for 5 minutes, 5 times a week for 4 weeks, at a pulse repetition rate of 20 Hz, with a peak compressive pressure of approximately 3 MPa. No abnormalities were detected in the carotid artery or surrounding tissue. No adverse effects were observed in the area where photoacoustic waves were applied. [Figure 6]Figure 6 is a schematic, not-to-scale, cross-sectional view of a tumor priming device according to some embodiments of the present disclosure, including a pulsed laser system (1) with a control system (6) that limits the number of laser pulses generated, and a light guide (2) that directs the laser pulses to an optical pressure transducer (3). A tumor-localizing support structure (4) positions the optical pressure transducer (3) near a selected region of a solid tumor (5), the tumor growing in the center of healthy tissue (7). Each pressure pulse generated by the optical pressure transducer through absorption of one laser pulse passes through a small (less than 3 cm) tissue or acoustic coupling medium path and then traverses at least a portion of the tumor mass. [Figure 7] 7 is a schematic, not-to-scale, cross-sectional view of a tumor priming device according to some embodiments of the present disclosure for generating pressure pulses near a solid tumor using an endoscope, including a pulsed laser system (1) with a control system (6) that limits the number of laser pulses generated, and a light guide (2) that directs the laser pulses to an optical pressure transducer (3). The tumor localization support structure (4) is an endoscope that positions the optical pressure transducer (3) less than 3 cm from a selected area of ​​a solid tumor (5) growing in the center of healthy abdominal tissue. Each pressure pulse is directed to the optical pressure transducer, where it is generated and passes through the intestinal wall to the tumor. [Figure 8] 8 is a schematic, not-to-scale, cross-sectional view of a tumor priming device according to some embodiments of the present disclosure for generating pressure pulses near a solid tumor using a catheter, including a pulsed laser system (1) with a control system (6) that limits the number of laser pulses generated, and a light guide (2) that may have a lens at its distal end that directs the laser pulses to an optical pressure transducer (3). The tumor localization support structure (4) is a urinary catheter with a balloon (8) that positions the optical pressure transducer (3) less than 3 cm from a selected area of ​​a solid tumor (5) growing in the bladder. Each pressure pulse is directed to the optical pressure transducer, where it is generated, and passes through the bladder to the tumor. [Figure 9] Figure 9 shows Kaplan-Meier plots of BALB / c mice bearing orthotopic 4T1 tumors. The plots show the control group (dash-dotted line, no priming and no treatment), the group with tumors exposed to photoacoustic waves (dashed line, priming and no treatment), the group treated with intraperitoneal administration of aCTLA4 (solid line, no priming and treatment), and the group with tumors exposed to photoacoustic waves and treated with intraperitoneal administration of aCTLA4 (dotted line, priming and treatment). Day 0 was the day treatment began, and tumors were selected when their longest diameter was at least 3 mm. Mice were euthanized when their longest diameter reached 12 mm. DETAILED DESCRIPTION OF THE INVENTION

[0051] For purposes of this disclosure, the following definitions apply:

[0052] The term "therapeutic agent" refers to a small molecule drug, biological agent, or immune cell that can be used to treat tumors, including chemotherapeutic agents, radiosensitizers, photosensitizers, nanoparticles, senolytic agents, biologics, immunomodulatory agents, immune molecules, or CD8+ T cells activated by tumor antigens and not suppressed by negative regulators. A therapeutic agent can be an agent approved by a regulatory agency for tumor or cancer treatment, an agent in clinical trials prior to regulatory approval, or an agent under investigation for tumor or cancer treatment.

[0053] The term "small molecule drug" refers to organic compounds with a molecular weight of 1 kDa or less. This term includes compounds that have desirable pharmacological properties and can be administered orally or by injection. Small molecule drugs include cytostatic or cytotoxic drugs used in cancer chemotherapy.

[0054] The term "photosensitizer" refers to dyes, possibly conjugated to a targeting moiety, that have no detectable therapeutic effect in their electronic ground state but, when electronically excited, can trigger processes that ultimately lead to cell death, as demonstrated by the photogeneration of reactive oxygen species in photodynamic therapy and photoimmunotherapy of cancer.

[0055] The term "macromolecule" refers to an organic or bioorganic molecule with a molecular weight greater than 1 kDa, which may be a protein, a bioconjugate, an RNA molecule, or a DNA molecule, or a fragment of these molecules.

[0056] The term "biologics" or "biological therapeutics" refers to a diverse group of pharmaceutical products including vaccines, growth factors, immunomodulators, monoclonal antibodies, and human blood and plasma-derived products. This definition specifically includes proteins purified from live culture systems or blood.

[0057] The term "immunomodulatory agent" refers to checkpoint inhibitors, costimulators of immune pathways, antibodies targeting immune cell antigens and / or cancer antigens, and cell therapy approaches (e.g., adoptive cell transfer with genetically engineered receptors such as chimeric antigen receptor therapy), and includes immunomodulatory agents such as ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, cemiplimab, dostallimab, tislelizumab, leratolimab, toripalimab, camrelizumab, and sintilimab, among others.

[0058] The term "nanomedicine" refers to a nanoparticulate drug carrier system in which, according to a number size distribution, 50% or more of the particles have a size ranging from 1 nm to 500 nm in one or more external dimensions, and which contains small molecule drugs, photosensitizers, or macromolecules.

[0059] "Tumor priming" refers to the normalization of blood vessels and reduction of solid state in solid tumors, including the subsequent or simultaneous reduction of interstitial fluid pressure within the solid tumor, facilitating the penetration of therapeutic agents into the solid tumor.

[0060] The term "piezoelectric optical transducer" or "optical pressure transducer" refers to a material that substantially absorbs the light of a laser pulse and converts the absorbed optical energy into a pressure pulse.

[0061] The term "pressure pulse" refers to a disturbance that involves a temporary density change in the medium through which it propagates. This definition of "pressure pulse" explicitly includes both shock waves and photoacoustic waves, which are also collectively referred to as "stress waves."

[0062] The term "pressure pulse tumor priming" refers to tumor priming with stress waves, which can be photoacoustic waves and / or shock waves.

[0063] The words "include," "includes," "includes," "including," "having," and variations thereof mean "including but not limited to."

[0064] The word "exemplary" is used herein to mean "serving as an example, instance, or illustration." Any embodiment described as "exemplary" is not necessarily to be construed as preferred or advantageous over other embodiments and / or to exclude the incorporation of features from other embodiments.

[0065] Throughout this application, various embodiments of the present disclosure may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the present disclosure. Accordingly, the description of a range should be considered to specifically disclose all possible subranges and individual numerical values ​​within that range. For example, a description of a range of 1 to 6 should be considered to specifically disclose subranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., as well as individual numerical values ​​within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

[0066] Whenever a range of values ​​is given herein, it is meant to include any recited value (fractional or integer) within that range. The phrases "a range between a first recited value and a second recited value" and "a range 'from' a first recited value to a second recited value" are used interchangeably herein and are meant to include the first and second recited values, and all fractional and integer values ​​therebetween.

[0067] The term "treatment" as used herein encompasses alleviating at least one symptom of, reducing the severity of, or inhibiting the progression of, a disease, disorder, or condition. Treatment does not necessarily mean that the disease, disorder, or condition is completely cured. To be an effective treatment, a useful composition herein need only reduce the severity of the disease, disorder, or condition, reduce the severity of symptoms associated therewith, or improve the quality of life of the patient or subject.

[0068] The term "subject" as used herein refers to animals, more particularly non-human mammals, humans, and human organisms. Non-human animal subjects also include prenatal forms of animals, such as embryos or fetuses. Non-limiting examples of non-human animals include horses, cows, camels, goats, sheep, dogs, cats, non-human primates, mice, rats, rabbits, hamsters, guinea pigs, and pigs. In one embodiment, the subject is a human.

[0069] It is understood that certain features of the present disclosure, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the present disclosure, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination, or in any other suitably described embodiment of the present disclosure. Certain features described in the context of various embodiments are not considered essential features of those embodiments, unless the embodiment cannot function without those elements.

[0070] In some embodiments, the present disclosure provides devices and methods for priming solid tumors with pressure pulses generated when a piezoelectric optical material absorbs a laser pulse, thereby improving the therapeutic efficacy of therapeutic agent administration.

[0071] Prior to this disclosure, it was completely unexpected that pressure pulses with spatial-peak time-average intensities below the power safety limits set by the FDA for diagnostic ultrasound could alter the tumor microenvironment (TME), even though they are well tolerated by normal tissue. Example 1 illustrates a typical stress wave generated by the absorption of a laser pulse in an optical (photovoltaic) transducer. Examples 2 and 3 demonstrate that such stress waves are safe for cells in vitro and tissues in vivo. The fact that these stress wave pressure pulses have no detectable effect on normal tissue is consistent with the fact that their spatial-peak time-average intensities are below the power safety limits set by the FDA for diagnostic ultrasound. The pressure pulses are completely safe and do not alter normal tissue. However, as shown in Example 4, exposing solid tumors to pressure pulses results in priming of the solid tumor and an enhanced response to anticancer therapeutic agents. One skilled in the art could not have predicted that pressure pulses well tolerated by normal tissue would be so effective in priming solid tumors.

[0072] According to some embodiments, the present disclosure provides an apparatus for tumor priming with pressure pulses, the apparatus including: a pulsed laser system; a light guide configured to guide laser pulses to one or more optical pressure transducers; one or more optical pressure transducers configured to absorb laser pulses from the pulsed laser system and generate pressure pulses; a tumor positioning support structure configured to couple the one or more optical pressure transducers to selected regions of a solid tumor; and a control system configured to limit exposure of the solid tumor to the pressure pulses.

[0073] In some embodiments, the pulse repetition rate of the pulsed laser system is between 0.1 Hz and 100 Hz.

[0074] In some embodiments, the device comprises a control system configured to limit exposure of the solid tumor to the pressure pulse for a period of time ranging from 1 second (s) to 60 minutes (min).

[0075] In some embodiments, the light guide configured to guide the laser pulses to the one or more optical pressure transducers comprises one or more optical fibers or light pipes.

[0076] In some embodiments, the light guide comprises a mirror, a lens, a prism, a diffuser, or a polarizer, or any combination thereof.

[0077] In some embodiments, the tumor localization support structure is configured to couple one or more optical pressure transducers to an acoustic coupling element disposed between the transducers and the surface of the solid tumor.

[0078] In some embodiments, the pressure pulses are photoacoustic waves. In some embodiments, the pressure pulses are shock waves. Priming a solid tumor with a pressure pulse comprises exposing the solid tumor to one or more pressure pulses.

[0079] The photoacoustic wave reaches a maximum compressive pressure p max It is a high-pressure ultrasonic pulse with a maximum instantaneous intensity of 15 MPa and a frequency exceeding 100 MHz.

[0080] I = p max 2 / (ρ v),

[0081] where ρ is the density of the medium (water: ρ = 997 kg / m 3 ), v is the speed of sound in the same medium (water: v = 1480 m / s), the maximum instantaneous intensity of such a photoacoustic wave is significantly higher: I = 15 kW / cm 2However, because the pulse duration of photoacoustic waves is similar to that of laser pulses, and the repetition rate of laser pulses with mJ energy is typically a few Hertz, maximum intensity is reached for only a small fraction of the time. For example, a laser pulse operating at 10 Hz with a 10 ns duration has a duty cycle of 10 -7 This means that the maximum instantaneous intensity is 15kW / cm 2 Even with this, the spatial-peak temporal-average intensity is only ISPTA = 1.5 mW / cm². Such photoacoustic waves are safe and exceed the FDA power limit for diagnostic ultrasound of 0.72 W / cm². 2 They do not produce cavitation and their effects are primarily mechanical. To approach the FDA safety limits for diagnostic ultrasound, pulse durations of 500 ns and pulse repetition rates of 100 Hz are required, with duty cycles of 5x10 -5 need to be reached.

[0082] Shock waves share with photoacoustic waves the fact that they carry temporary density changes within the material through which they propagate. The difference is that shock waves propagate at a speed higher than the local speed of sound within the material. Herein, photoacoustic waves and shock waves are collectively referred to as "pressure pulses." Those skilled in the art will recognize that shock waves can be generated by a variety of processes, including explosions, projectiles impacting a surface, objects moving at ultrasonic speeds, or intense pulsed lasers causing target ablation. In the context of the present disclosure, the generation of shock waves by intense laser pulses is of particular interest. The laser fluence rate (W / m) required to generate a plasma at a given target and consequently generate shock waves is 2The pressure (in units) is higher than that required for thermoelastic expansion of the target and, consequently, for generating photoacoustic waves. This means that shock waves generated by pulsed lasers typically result in higher peak pressures than photoacoustic waves using the same material. Nevertheless, the peak intensities of both shock waves and photoacoustic waves are achieved within a very short time. Therefore, low pulse repetition rates (<100 Hz) result in very low duty cycles, and pressure pulses of up to 100 MPa can be used without significant damage to tissue when the pressure pulses are generated by laser pulses with nanosecond durations.

[0083] In some embodiments of the present disclosure, superficial solid tumors can be exposed to pressure pulses by placing a laser pulse-absorbing material directly over the tumor or over the skin layer covering the tumor, thereby providing good acoustic coupling with the skin and tumor. A laser pulse is directed at the material, the energy of the laser pulse is absorbed by the material, and either photoacoustic waves or shock waves are generated in the material and transmitted through the material to the skin and tumor. Good acoustic coupling can be achieved by properly matching the acoustic impedance of the material to human tissue, and can be improved by using an acoustic coupling gel layer. The transition from thermoelastic expansion, which generates photoacoustic waves, to ablation, which generates shock waves, depends primarily on the ablation threshold of the material, the energy of the laser pulse, and the size of the irradiated area. See Example 1 in the Examples section of this disclosure, which describes a method for generating and characterizing photoacoustic waves.

[0084] In many clinical situations, solid tumors are not superficial, i.e., located more than 3 cm below the body surface. In such situations, stress waves generated at the body surface may be strongly attenuated by healthy tissue before reaching the tumor mass, resulting in a loss of tumor priming efficacy. According to the FDA, ultrasound attenuation in tissue can be calculated using an attenuation coefficient of 0.3 dB / (cmMHz). This means that at a distance of 3 cm from a 3.3 MHz transducer, the time-averaged intensity after attenuation is 3 dB (i.e., half) of the value measured in water. However, higher frequencies have a stronger effect on tumor priming; applying the same attenuation coefficient to a 33 MHz frequency at 3 cm results in a time-averaged intensity after attenuation of 30 dB (i.e., 0.001 times) the value measured in water. This indicates that to significantly expose solid tumors to stress waves, the optical pressure transducer must be placed less than 3 cm from the surface of the solid tumor. This disclosure is based, in part, on the surprising discovery that normal cells and healthy tissue are not affected by stress waves. See Examples 2 and 3 in the Examples section of this disclosure. These indicate that normal cells and healthy tissues are not affected by stress waves.

[0085] In some embodiments, even if a solid tumor is not superficial, it may be possible to access the tumor mass through a natural body cavity using endoscopic methods. Minimally invasive procedures can be used to generate stress waves near solid tumors in the gastrointestinal tract, respiratory system, urinary system, or female reproductive system. In these locations, endoscopes have channels that allow for the insertion of optical fibers, allowing stress waves to be generated near solid tumors. Alternatively, small incisions less than 2 cm long can be made in procedures such as laparoscopy and thoracoscopy to allow optical fiber access to normally closed body cavities. Furthermore, optical fibers can be inserted into catheters to reach many target locations within the human body.

[0086] In some embodiments of the present disclosure, the use of optical fibers allows for the delivery of laser light to the vicinity of solid tumors. Non-limiting examples of solid tumors accessible by optical fibers include gastric cancer, intestinal cancer, lung cancer, breast cancer, uterine cancer, esophageal cancer, ovarian cancer, pancreatic cancer, pharyngeal cancer, sarcoma, liver cancer, urinary tract and bladder cancer, maxillary cancer, bile duct cancer, head and neck cancer, tongue cancer, brain tumor, skin cancer, malignant goiter, prostate cancer, colon cancer, parotid cancer, and kidney cancer.

[0087] In some embodiments, when an optical fiber is used to deliver laser light to a solid tumor, the laser light can be directed to the proximal end of the optical fiber, and a piezoelectric-optical transducer can be coupled to its distal end, which is located near the solid tumor. The piezoelectric-optical transducer absorbs most of the intensity of the laser pulse, generating stress waves. Each time a laser pulse is absorbed by the piezoelectric-optical transducer, a stress wave is generated. Stress waves can be generated by thermoelastic expansion of the piezoelectric-optical transducer under conditions of thermal and stress confinement, in which case the stress wave is a photoacoustic wave. Stress waves can also be generated by ablation of the piezoelectric-optical transducer, which involves the removal or destruction of some material in the transducer, in which case the stress wave is a shock wave. In either case, the stress wave propagates through the piezoelectric-optical transducer from the side where the laser pulse was absorbed to the other side, and is then transmitted to tissue near the solid tumor or directly to the solid tumor.

[0088] According to the present disclosure, piezoelectric optical transducers can be made from a variety of dyes or pigments, but to generate pressure pulses with high peak compression amplitudes, the dye or pigment must have a high absorption coefficient (μ) at the wavelength of the laser pulse and must convert the absorbed optical energy into thermal energy quickly and efficiently. Furthermore, if the intention is to generate stress waves that are photoacoustic waves, the dye or pigment should preferably be incorporated into a material with a high Gruneisen parameter (Γ > 0.5), since the peak pressure of a photoacoustic wave is given by:

[0089] p0= Γ μ F,

[0090] where F is the local light fluence. If the intention is to generate stress waves that are shock waves, the dye or pigment should preferably be incorporated into a material with a low ablation threshold. This is the case, for example, with poly(ethylene terephthalate), polyimides, and triazene polymers. These polymers have a fluence of 200 mJ / cm. 2 This can be used to produce piezo-optical materials that undergo ablation accompanied by the generation of shock waves at laser fluences below 1000 .mu.m.

[0091] Non-limiting examples of dyes or pigments that can be used to fabricate piezoelectric optical materials according to the present disclosure include ortho-hydroxybenzophenone and similar molecules that undergo ultrafast photoinduced intramolecular proton or hydrogen atom transfer and rapidly return to their original ground state; meso-tetraphenylporphyrin, a MnIII complex, and other paramagnetic complexes with ultrafast metal-ligand and / or ligand-metal charge transfer relaxation processes; complexes with charge transfer bands that return to their ground state via ultrafast charge recombination; β-carotene and other systems that rapidly decay to their ground state through a conical intersection; graphite or carbon nanoparticles, carbon nanotubes, or carbon soot, which are capable of ultrafast transfer of electronic energy to phonon modes following cooling on subnanosecond timescales; semiconductor materials with short-lived transition states; or other materials or mixtures of materials with ultrafast nonradiative relaxation processes. In addition to the conversion of light energy to thermal energy during ablation, structural volume changes in dyes and pigments can also occur, potentially contributing to increased stress wave intensity.

[0092] In some embodiments, the optical pressure transducer includes a laser light absorption system and a material with a Gruneisen parameter higher than 0.5, and each pressure pulse is a photoacoustic wavefront.

[0093] Non-limiting examples of materials with a high Gruneisen parameter (G > 0.5) include polymers (polydimethylsiloxane, polystyrene, polyamide, poly(vinyl chloride), polyethylene, polyacrylonitrile, poly(ethylene terephthalate), polychloroprene, parylene), metal films, glass, and layered materials containing them. Such materials can absorb the light of a laser pulse over a very short optical path, or can be engineered to incorporate dyes or pigments that absorb the light of a laser pulse, making them very useful for manufacturing piezoelectric optical materials that function with endoscopes or catheters.

[0094] In some embodiments, the optical pressure transducer comprises a laser light absorption system and an ablation threshold of 200 mJ / cm 2 Each pressure pulse is a shock wave front.

[0095] In some embodiments, piezoelectric-optical transducers made from dyes or pigments and materials with high Gruneisen parameters or low ablation thresholds can take a variety of shapes. Given that dyes or pigments must have high absorption coefficients, piezoelectric-optical transducers can absorb a large portion of the laser pulse with an optical path of less than 200 μm, preferably less than 100 μm, and most preferably less than 50 μm. Given the extremely small thickness of piezoelectric-optical transducers, they can be used to cover the distal end of an optical fiber.

[0096] In some embodiments, the peak compressive pressure of the stress wave ranges from 0.1 MPa to 100 MPa.

[0097] In some embodiments, 90% of each pressure pulse lasts between 0.1 ns and 500 ns.

[0098] 6, 7 and 8 show various embodiments of devices according to the present disclosure in which a piezoelectric optical transducer is coupled to the distal end of an optical fiber.

[0099] In some embodiments, the optical fiber is carried by a tumor localization support structure, which directs the laser pulses and optical pressure transducer to a solid tumor less than 3 cm from the body surface (FIG. 6). In another embodiment, the optical fiber is inserted into an endoscope, which functions as the tumor localization support structure, and at its distal end, the optical fiber is optically coupled to an optical diffuser, which is at least partially coated with a piezoelectric optical transducer and positioned within 3 cm of the solid tumor (FIG. 7). In some embodiments, the optical fiber is inserted with the aid of a balloon into a catheter, which functions as the tumor localization support structure, and at its distal end, the optical fiber has a lens that directs the laser pulse to a piezoelectric optical transducer positioned within 3 cm of the solid tumor (FIG. 8). In another embodiment, the optical fiber is coupled to an optical diffuser, which is coated with a piezoelectric optical transducer and inserted into the solid tumor, and the system that perforates the tumor and inserts the optical pressure transducer is the tumor localization support structure, and the structure has a sharp tip.

[0100] In some embodiments, the tumor localization support structure is an endoscope and the light guide is one or more optical fibers configured to carry laser light from a light source through the endoscope to one or more optical pressure transducers at the distal end of the optical fiber.

[0101] In some embodiments, the endoscope is configured for insertion into a hollow organ through a natural body cavity opening or a body incision less than 2 cm in length.

[0102] In some embodiments, the tumor localization support structure includes a sharpened tip configured to allow insertion of one or more optical pressure transducers into a solid tumor.

[0103] In some embodiments, the catheter is configured for insertion into a body cavity, duct, blood vessel, brain, skin, or fatty tissue.

[0104] In some embodiments, the tumor localization support structure includes a sharpened end configured to allow insertion of one or more optical pressure transducers into a solid tumor.

[0105] Tumor priming using pressure pulses involves placing a photovoltaic material within 3 cm of a solid tumor, ensuring that the path between the photovoltaic material and the solid tumor is filled with a medium capable of transmitting the pressure pulse, irradiating the photovoltaic material with a laser pulse that generates a peak compressive pressure of 0.1 to 100 MPa within the photovoltaic material, and directing such pressure pulses at at least a portion of the solid tumor for a time period ranging from 1 second to 1 hour. The laser pulse may have a femtosecond, picosecond, or nanosecond duration (full width at half maximum). Preferably, the laser pulse duration should be less than 500 nanoseconds, because thermal confinement conditions are more easily met under these conditions, and 90% of pressure pulses last less than 500 ns.

[0106] In some embodiments, the solid tumor can be exposed to the pressure pulse for a short time (e.g., 1 second), a long time (e.g., 1 hour), or an intermediate time period. The solid tumor can be exposed to the pressure pulse before, during, or after administration of the therapeutic agent, and the timing can be adjusted depending on the plasma lifetime of the therapeutic agent.

[0107] The present disclosure is based, in part, on the discovery that for repeated administration of therapeutic agents and / or therapeutic agents with long plasma half-lives, exposure of solid tumors to pressure pulses can be performed several times, such as several times per day, several times per week, several times per month, or several times per year.

[0108] The present disclosure is based, in part, on the discovery that pressure pulse-mediated solid tumor priming, as described herein, promotes the penetration of therapeutic agents into solid tumors and enhances their response to therapy. Solid tumor priming by exposure to pressure pulses promotes the penetration of various therapeutic agents into solid tumors without affecting the delivery of the therapeutic agent to healthy host tissues or increasing host toxicity. This is particularly valuable for the delivery of small molecule drugs and macromolecules that bind extensively to plasma proteins, particularly biopharmaceuticals. The present disclosure is based, in part, on the discovery that pressure pulse-mediated solid tumor priming improves the therapeutic efficacy of immunotherapy, particularly when the therapeutic agent is a mAb used in ICB therapy. Pressure pulse-mediated solid tumor priming promotes the penetration of tumor antigen-specific T lymphocytes into tumors and their integration into the tumor microenvironment (TME), contributing to an enhanced tumor response to immunotherapy. The present disclosure is based, in part, on the unexpected discovery that pressure pulses, which are well tolerated by normal tissue, are effective for solid tumor priming.

[0109] In some embodiments, the present disclosure provides a method of treating a tumor in a subject. In some embodiments, the method includes administering to the subject (i) an amount of high-intensity photoacoustic waves and (ii) an amount of a therapeutic agent, wherein the amounts (i) and (ii) together are sufficient to treat a solid tumor, and the order of administration can be selected from (i) before (ii), (i) simultaneously with (ii), or (i) after (ii). In some embodiments, a method of sensitizing a tumor in a subject to an amount of anti-cancer therapy is disclosed, the method comprising administering to the subject an amount of pressure pulses effective to improve the response of the tumor in the subject to the amount of anti-cancer therapy administered to the subject before or during the course of the anti-cancer therapy. In some embodiments, the subject is a cancer patient.

[0110] In some embodiments, the present disclosure provides a method for treating a solid tumor in a subject suffering from cancer, the method comprising: pressure pulse tumor priming by exposing the solid tumor to one or more pressure pulses, the pressure pulses having a maximum compressive pressure of between 0.1 MPa and 100 MPa, with 90% of each pressure pulse lasting between 1 nanosecond and 500 nanoseconds; and administering to the subject one or more anti-cancer therapeutic agents, thereby treating the solid tumor in the subject suffering from cancer. In some embodiments, the subject suffering from cancer is a cancer patient.

[0111] In some embodiments, the step of pressure pulse tumor priming by exposing the solid tumor to one or more pressure pulses is performed before administration of one or more anti-cancer therapeutic agents to the subject, during administration of one or more anti-cancer therapeutic agents to the subject, or after administration of one or more anti-cancer therapeutic agents to the subject.

[0112] In some embodiments, the method further comprises repeating (i) pressure pulse tumor priming, (ii) administration of one or more anti-cancer therapeutics, or both, at least 1, 2, 3, 5, or as many times as necessary to alleviate symptoms in a subject suffering from cancer, at doses that improve the solid tumor's response to treatment.

[0113] In some embodiments, the anti-cancer therapeutic agent is selected from the group consisting of an inhibitor of an inhibitory checkpoint molecule, an activator of a stimulatory checkpoint molecule, an antibody, a cytokine, an interferon, an interleukin, a vaccine, an oncolytic virus, a chimeric antigen receptor T cell, and any combination thereof.

[0114] In some embodiments, the therapeutic agent is a biotherapeutic agent.

[0115] In some embodiments, the therapeutic agent is a monoclonal antibody (mAb) used in cancer treatment, or any combination of mAbs used in cancer treatment.

[0116] In some embodiments, the therapeutic agent is selected from ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, cemiplimab, dostallimab, tislelizumab, leratolimab, toripalimab, camrelizumab, sintilimab, or any combination thereof.

[0117] In some embodiments, the therapeutic agent is a cytostatic or cytotoxic drug bound to a plasma protein.

[0118] In some embodiments, the therapeutic agent is a macromolecule.

[0119] In some embodiments, the therapeutic agent is a nanomedicine.

[0120] According to some embodiments, the present disclosure provides a kit comprising the above-described device and the above-described anti-cancer therapeutic agent.

[0121] According to some embodiments, the present disclosure provides a system including at least one device as described above.

[0122] According to some embodiments, the present disclosure provides a device for treating solid tumors in cancer patients, the device comprising: means for generating pressure pulses having a peak compressive pressure of 0.1 MPa to 100 MPa, with 90% of each pressure pulse lasting between 1 ns and 500 ns. In some embodiments, the device is as described above.

[0123] Pressure pulse tumor priming therapy (PPTPT) combines the exposure of solid tumors to pressure pulses and the administration of anticancer therapeutic agents, as described above. Example 4 and Figure 9 demonstrate PPTPT in mice bearing orthotopic 4T1 breast cancer. 4T1 cells were inoculated into the mammary fat pad of BALB / c mice and allowed to grow to 3 mm in their longest diameter before the start of intervention. In this example, pressure pulse tumor priming consisted of exposing the orthotopic tumor to photoacoustic waves for 5 minutes on days 0 and 2. Treatment in this example consisted of intraperitoneal administration of anti-CTLA-4 mAb (aCTLA4) on days 0, 2, 6, and 10. The control group demonstrates that the survival of mice in the group receiving tumor priming alone (primed, no treatment) was not statistically different from that of the control group (no priming, no treatment). In the group receiving four doses of aCTLA4 (no priming, treatment), only one animal responded to treatment. In contrast, all animals in the group receiving PPTPT (primed, treated) responded to treatment.

[0124] The extraordinary results of photoacoustic tumor priming can be seen by comparing PPTPT with tumor priming using angiotensin receptor blockers

[17] . The median survival time of mice bearing orthotopic 4T1 tumors was 20 days when aCTLA4 and aPD1 were combined, but this was extended to 24 days when this combination was combined with tumor priming using angiotensin receptor blockers

[17] . Using the same animal model, i.e., mice bearing orthotopic 4T1 tumors, Example 4 demonstrates that median survival time was 21 days when animals were treated with aCTLA4, but was extended to 36 days when photoacoustic priming was combined with aCTLA4 treatment. Those skilled in the art would not have predicted that exposing orthotopic 4T1 tumors to photoacoustic waves for 5 minutes in two separate sessions would increase the median survival time of orthotopic 4T1 tumor-bearing mice by 15 days. This is particularly remarkable when considering that priming with angiotensin receptor blockers only increased median survival by 4 days. Orthotopic 4T1 tumors are widely recognized to be extremely difficult to treat, and it is completely unexpected that two sessions of local exposure lasting just 5 minutes increased the immunotherapy response in all mice to tumors exposed to harmless photoacoustic waves.

[0125] This disclosure is based, in part, on the discovery that PPTPT is a novel and surprisingly effective approach for enhancing the response of solid tumors to therapeutic agents. High-intensity broadband stress waves exert mechanical forces at the microscopic level and can remodel the TME. As shown in Example 1, a peak pressure of approximately 7 MPa for a pressure wave with a relevant frequency of approximately 20 MHz corresponds to a 50 bar change in 10 nanoseconds, or a 50 bar change in 15 μm, considering the speed of sound propagation in tissue. Dramatic pressure changes occur on the scale of cells. This disclosure is based, in part, on the discovery that cells can tolerate these high pressures, as shown in Example 2, and that normal tissue shows no adverse effects, as shown in Example 3. However, as shown in Example 4, the mechanical forces exerted by such pressure pulses on the TME enable micromechanical priming of solid tumors.

[0126] Various embodiments and aspects of the present disclosure as delineated above and delineated in the claims section below find experimental support in the following examples.

[0127] <References> [1] I. Mellman, G. Coukos, G. Dranoff, Cancer immunotherapy comes of age, Nature, 480 (2011) 480-489. [2] JM Kirkwood, LH Butterfield, AA Tarhini, H. Zarour, P. Kalinski, S. Ferrone, Immunotherapy of Cancer in 2012, CA Cancer J. Clin., 62 (2012) 309-335. [3] SJ Casak, L. Marcus, L. Fashoyin-Aje, S. Mushti, J. Cheng, Y.-L. Shen, WF Pierce, L. Her, KB Goldberg, MR Theoret, PG Kluetz, R. Pazdur, SJ Lemery, FDA Approval Summary: Pembrolizumab for the First-line Treatment of Patients with MSI-H / dMMR Advanced Unresectable or Metastatic Colorectal Carcinoma, Clin. Cancer Res., 27 (2021) 4680-4684. [4] JD Twomey, B. Zhang, Cancer Immunotherapy Update: FDA-Approved Checkpoint Inhibitors and Companion Diagnostics, AAPS J., 23 (2021) 39. [5] A. Haslam, V. Prasad, Estimation of the Percentage of US Patients With Cancer Who Are Eligible for and Respond to Checkpoint Inhibitor Immunotherapy Drugs, JAMA Network Open, 2 (2019) e192535. [6] J.M. Fritz, M.J. Lenardo, Development of immune checkpoint therapy for cancer, J. Exp. Med., 216 (2019) 1244-1254. [7] A.D. Waldman, J.M. Fritz, M.J. Lenardo, A guide to cancer immunotherapy: from T cell basic science to clinical practice, Nat. Rev. Immunol., 20 (2020) 651-668. [8] A.I. Minchinton, I.F. Tannock, Drug penetration in solid tumours, Nat. Rev.Cancer, 6 (2006) 583-592. [9] J.T. Ryman, B. Meibohm, Parmacokinetics of monoclonal antibodies, CPT Pharmacometrics Syst. Pharmacol., 6 (2017) 576-588.

[10] M. Centanni, D.J.A.R. Moes, I.F. Troconiz, J. Ciccolini, J.G.C. van Hasselt, Clinical Pharmacokinetics and Pharmacodynamics of Immune Checkpoint Inhibitors, Clin. Pharmacokinet., 58 (2019) 835-857.

[11] Y. Matsumura, H. Maeda, A New Concept for Macromolecular Therapeutics in Cancer Chemotherapy: Mechanism of Tumoritropic Accumulation of Proteins and the Antitumor Agent Smancs, Cancer Res., 46 (1986) 6387-6392.

[12] Y.H. Bae, K. Park, Targeted drug delivery to tumors: Myths, reality and possibility, J. Control. Release, 153 (2011) 198-295.

[13] Y. Huang, J. Yuan, E. Righi, W.S. Kamoun, M. Ancukiewicz, J. Nezivar, M. Santosuosso, J.D. Martin, M.R. Martin, F. Vianello, P. Leblanc, L.L. Munn, P. Huang, D.G. Duda, D. Fukumura, R.K. Jain, M.C. Poznansky, Vascular normalizing doses of antiangiogenic treatment reprogram the immunosuppressive tumor microenvironment and enhance immunotherapy, Proc. Natl. Acad. Sci. USA, 109 (2012) 17561-17566.

[14] I.A. Khawar, J.H. Kim, H.-J. Kuh, Improving drug delivery to solid tumors: Priming the tumor microenvironment, J. Control. Release, 201 (2015) 78-89.

[15] T. Stylianopoulos, J.D. Martin, V.P. Chauhan, S.R. Jain, B. Diop-Frimpong, N. Bardeesy, B.L. Smith, C.R. Ferrone, F.J. Hornicek, Y. Boucher, L.L. Munn, R.K. Jain, Causes, consequences, and remedies for growth-induced solid stress in murine and human tumors, Proc. Natl. Acad. Sci. USA, 109 (2012) 15101-15108.

[16] H. Raskov, A. Orhan, J.P. Christensen, I. Gogenur, Cytotoxic CD8+ T cells in cancer immunotherapy, Br. J. Cancer, 124 (2021) 359-367.

[17] V.P. Chauhan, Y.X. Ivy X. Chen, R. Tong, M.R. Ng, J.D. Martin, K. Naxerova, M.W. Wu, P. Huang, Y. Boucher, D.S. Kohane, R. Langer, R.K. Jain, Reprogramming the microenvironment with tumor-selective angiotensin blockers enhances cancer immunotherapy, Proc. Natl. Acad. Sci. USA, 116 (2019) 10674-10680.

[18] H.-C. Huang, I. Rizvi, J. Liu, S. Anbil, A. Kalra, H. Lee, Y. Baglo, N. Paz, D. Hayden, S. Pereira, BW Pogue, J. Fitzgerald, T. Hasan, Photodynamic Priming Mitigates Chemotherapeutic Selection Pressures and Improves Drug Delivery, Cancer Res., 78 (2018) 558-571.

[19] C. Guha, S. Barry, F. Macian, Low intensity focused ultrasound for treating cancer and metastasis, in: US Patent (Ed.), Montefiore Medical Center, USA, 2021; US ​​10,974,077.

[0128] The following examples, together with the above description, illustrate some embodiments of the present disclosure in a non-limiting fashion.

[0129] Example 1 Photoacoustic wave generation with a peak pressure of 10 MPa

[0130] Carbon nanoparticles are a very useful optical absorption system because they strongly absorb light over a wide range of wavelengths, from ultraviolet to visible to infrared. Because carbon nanoparticles are difficult to disperse in solution, 160 mg of carbon nanoparticles produced as candle soot were added to 5 mL of toluene and sonicated for 5 minutes at 60 MHz using a tip sonicator. Immediately after mechanical sonication, 2 mg of polystyrene was added to the suspension and heated to 60 °C in a water bath. Polystyrene has a high Gruneisen parameter (G ≒ 0.7), making it very convenient for fabricating thin piezoelectric optical transducers. Polystyrene films containing dispersed carbon nanoparticles were fabricated using a mechanical applicator (Elcometer) and allowed to dry overnight to allow the remaining solvent to evaporate.

[0131] A piezoelectric optical transducer was also fabricated by depositing carbon nanoparticles from the combustion of a paraffin lamp onto a borosilicate glass window. This glass window was exposed to a direct flame for 2 minutes to collect the carbon soot. The thin carbon soot layer deposited on the window was then covered with 0.1 mL of polydimethylsiloxane (PDMS) and treated under vacuum for 10 minutes to remove air bubbles. Next, a 100 g weight was placed on top of the system (glass + soot + PDMS) to form a thin layer of PDMS, which was then subjected to vacuum for another 10 minutes to remove excess air trapped within the system. Finally, the complete assembled system was heated overnight in an oven at 50 °C to achieve complete curing of the PDMS.

[0132] Piezoelectric optical transducers fabricated with carbon nanoparticles and polystyrene or PDMS as described above were investigated with 1064 nm pulsed laser excitation to characterize the pressure pulses they could generate. Laser excitation was performed using a Nd:YAG laser (Monfort M-NANO) with nanosecond pulses to generate photoacoustic waves. Two types of ultrasonic measurements were performed. Absolute pressure was measured using a 0.2 mm needle-type hydrophone (Precision Acoustics, model NH0200) calibrated for the 1-30 MHz range. Ultrasonic frequency distribution was investigated with a 225 MHz contact transducer (Panametrics / Olympus, model V2113). Figure 1A-B shows the ~60 mJ / cm measured with the hydrophone. 2 and ~250mJ / cm 2 Figure 2 shows the absolute pressure pulse obtained at a laser fluence of ~60 mJ / cm measured with a contact transducer. 2 4 shows the ultrasonic frequency distribution using a laser fluence of

[0133] <Example 2> Photoacoustic waves with a peak pressure of 10 MPa are not toxic to fibroblasts in vitro

[0134] Monolayer cultures of immortalized monkey fibroblast cell line (COS-7) were grown in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (Gibco) and 1% penicillin and streptomycin (Invitrogen) in a humidified atmosphere at 37°C with 5% CO2. COS-7 cells were first seeded into 12-well plates at a density of 30,000 cells / well in 2 mL of medium. After 24 hours, the medium was replaced to allow rapid cell growth. 48 hours after seeding, the medium was removed and 300 μL of fresh culture medium was added. Next, the piezoelectric optical material described in Example 1 was immersed in the culture medium and positioned within 3 mm of the surface of the COS-7 cell monolayer. The cells were then irradiated with an Nd:YAG laser (Monfort M-NANO) at a laser repetition rate of 6 Hz or 20 Hz and a laser fluence of approximately 60 mJ / cm. 2 Cells were exposed to photoacoustic waves for 5 minutes (5 min) or 10 minutes (10 min) at 100°C. Cell viability was measured using the Alamar Blue® assay 24 hours after exposure to photoacoustic waves.

[0135] Example 3 Exposing healthy rat tissue to photoacoustic waves for five minutes a day, five days a week for four weeks did not cause any adverse effects.

[0136] The Portuguese Animal Health Agency approved the animal experiments (DGAV authorization 0420 / 000 / 000 / 2011). Male Sprague-Dawley albino rats (Charles River Laboratories, Barcelona, ​​Spain) were used in this study. The hair around the rat's neck was removed, and a circle was drawn over the area where stress waves were to be applied. Exposure to stress waves was carried out 5 days a week for 4 weeks. During each exposure, stress waves were generated at 20 Hz for 5 minutes using a piezoelectric-optical transducer made of carbon nanoparticles and PDMS and a Monfort M-NANO Nd:YAG laser. Under the conditions used, the peak compressive pressure of each pulse was approximately 3 MPa. Acoustic coupling between the piezoelectric-optical transducer and the rat's neck was optimized using ultrasound gel (Eco Supergel). The carotid arteries were imaged using magnetic resonance imaging (MRI) (Figure 4). Histological examination of neck sections was performed at the end of the experiment (Figure 5).

[0137] Example 4 Pressure Pulse Tumor Priming Therapy

[0138] The Portuguese Animal Health Authority approved the animal experiments (DGAV authorization 0420 / 000 / 000 / 2011). 4T1 cells (ATCC CRL-2539) were cultured in Dulbecco's modified Eagle's medium (DMEM) (Sigma-Aldrich, Saint-Louis, MO, USA) supplemented with 10% (v / v) heat-inactivated fetal bovine serum (GIBCO®, Life Technologies, Bleiswijk, The Netherlands), 100 U / mL penicillin, and 100 ng / mL streptomycin (Invitrogen®, Thermo Fisher Scientific, Grand Island, NY, USA). Tumors were established by orthotopic injection of 20,000 4T1 cells into the right mammary gland of approximately 8-12 week-old (20 g) female BALB / c mice.

[0139] Prior to tumor priming, the mice were depilated from their abdominal region, particularly the mammary glands where the tumor was inoculated. The piezoelectric transducer used was fabricated from carbon nanoparticles and polydimethylsiloxane. The piezoelectric transducer was placed on top of the tumor, and acoustic coupling was improved by placing a layer of acoustic coupling gel between the tumor and the piezoelectric transducer. Photoacoustic waves were generated by directing laser pulses from a Monfort M-NANO Nd:YAG laser at a laser repetition rate of 20 Hz onto the piezoelectric transducer. Under the conditions used, the peak compressive pressure of each pulse was approximately 6.5 MPa.

[0140] This protocol used four study groups, each consisting of 4–5 mice: (i) a control group with orthotopic tumors, without tumor priming or treatment; (ii) a priming control group with orthotopic tumors, tumor priming by pressure pulse, and no treatment; (iii) an anti-mouse CTLA4 treatment group with orthotopic tumors, without tumor priming, and treatment with in vivo mAb anti-mouse CTLA4 (CD152); and (iv) a tumor priming treatment group with orthotopic tumors, tumor priming by pressure pulse, and treatment with in vivo mAb anti-mouse CTLA4 (CD152). Day 0 (zero) was defined as the first treatment day; tumors in all groups were approximately 3 mm in diameter. Day 0 corresponds to 8 days after orthotopic tumor inoculation.

[0141] Group (i) received no tumor priming or treatment, and orthotopic tumors were allowed to grow naturally. Group (ii) underwent tumor priming by exposing tumors to photoacoustic waves for 5 minutes on days 0 and 2. Group (iii) was treated with InVivo mAb anti-mouse CTLA4 (CD152) (BioCell, Lebanon, NH, USA) via intraperitoneal injection on days 0, 2, 6, and 10. Group (iv) underwent the same treatment protocol as group (iii), but additionally underwent tumor priming 10 minutes after antibody administration on days 0 and 2, as in group (ii). Tumors were measured twice weekly with a caliper, and animals were euthanized when tumors reached a diameter of 12 mm. Figure 9 shows the survival of mice to this endpoint.

[0142] While this disclosure has been described in conjunction with specific embodiments thereof, it is evident that many alternatives, modifications, and variations will be apparent to those skilled in the art. Accordingly, the present disclosure is intended to embrace all such alternatives, modifications, and variations that fall within the spirit and broad scope of the appended claims.

[0143] All publications, patents, and patent applications mentioned in this specification are incorporated herein by reference in their entirety to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated herein by reference. Furthermore, citation or identification of any reference in this application should not be construed as an admission that such reference is available as prior art to the present disclosure. To the extent headings are used, they should not be construed as necessarily limiting.

Claims

1. 1. A device for tumor priming by pressure pulses, comprising: - Pulsed laser systems with pulse repetition rates between 0.1Hz and 100Hz; - an optical guide configured to direct laser pulses to one or more optical pressure transducers; one or more optical pressure transducers configured to absorb laser pulses from the pulsed laser system and generate pressure pulses, the pressure pulses having a peak compression pressure between 0.1 MPa and 100 MPa and 90% of each pressure pulse lasting between 0.1 ns and 500 ns; a tumor localization support structure configured to couple one or more optical pressure transducers to a selected region of a solid tumor at a distance of less than 3 cm from said region; a control system configured to limit exposure of the solid tumor to said pressure pulse for a period of between 1 second and 60 minutes.

2. 10. The device of claim 1, wherein the light guide comprises one or more optical fibers or light pipes.

3. 10. The device of claim 1, wherein the light guide comprises a mirror, a lens, a prism, a diffuser, a polarizer, or any combination thereof.

4. 10. The device of claim 1, wherein the optical pressure transducer comprises a laser light absorption system and a material having a Gruneisen parameter higher than 0.5, and each pressure pulse is a wavefront of a photoacoustic wave.

5. 10. The device of claim 1, wherein the optical pressure transducer comprises a laser light absorption system and an ablation threshold of 200 mJ / cm 2 A device containing less than 1000 psi of material, where each pressure pulse is the wavefront of a shock wave.

6. 10. The device of claim 1, wherein the tumor localization support structure is configured to hold one or more optical pressure transducers with an acoustic coupling element positioned between the transducers and a surface of a solid tumor.

7. 10. The device of claim 1, wherein the tumor localization support structure is an endoscope and the light guide is one or more optical fibers configured to carry laser light from a light source through the endoscope to one or more optical pressure transducers at the distal end of the optical fibers.

8. 8. The device of claim 7, wherein the endoscope is configured for insertion into a hollow organ through a natural body cavity opening or a body incision less than 2 cm in length.

9. 10. The device of claim 1, wherein the tumor location support structure is a catheter and the light guide is one or more optical fibers configured to carry laser light from a light source through the catheter to one or more optical pressure transducers at the distal end of the optical fiber.

10. 10. The device of claim 9, wherein the catheter is configured for insertion into a body cavity, duct, blood vessel, brain, skin, or fatty tissue.

11. 10. The device of claim 1, wherein the tumor localization support structure includes a sharpened end that allows one or more optical pressure transducers to be inserted into the solid tumor.

12. 1. A method of treating a solid tumor in a subject afflicted with cancer, comprising: - pressure pulse tumor priming of a solid tumor by exposing the solid tumor to one or more pressure pulses, the pressure pulses having a peak compressive pressure of 0.1 MPa to 100 MPa, with 90% of each pressure pulse lasting between 1 ns and 500 ns; and administering to said subject one or more anti-cancer therapeutic agents; Thereby, the method of treating said solid tumor in a subject suffering from cancer.

13. 13. The method of claim 12, wherein the pressure pulse priming of the solid tumor is performed with the device of claim 1.

14. 13. The method of claim 12, further comprising repeating the pressure pulse priming, the administration of the one or more anti-cancer therapeutic agents, or both, at least once at a dose that improves the therapeutic response of the solid tumor.

15. 13. The method of claim 12, wherein the anti-cancer therapeutic agent is selected from the group consisting of an inhibitor of an inhibitory checkpoint molecule, an activator of a stimulatory checkpoint molecule, an antibody, a cytokine, an interferon, an interleukin, a vaccine, an oncolytic virus, a chimeric antigen receptor T cell, and any combination thereof.

16. 13. The method of claim 12, wherein the therapeutic agent is a biological therapeutic agent.

17. 13. The method of claim 12, wherein the therapeutic agent is a monoclonal antibody (mAb) used in the treatment of cancer, or any combination of mAbs used in the treatment of cancer. Any combination of mAbs used in treatment.

18. 13. The method of claim 12, wherein the therapeutic agent is selected from ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, cemiplimab, dostallimab, tislelizumab, leratolimab, toripalimab, camrelizumab, sintilimab, or any combination thereof.

19. 13. The method of claim 12, wherein the therapeutic agent is a cytostatic or cytotoxic drug bound to a plasma protein.

20. 13. The method of claim 12, wherein the therapeutic agent is a macromolecule.

21. 13. The method of claim 12, wherein the therapeutic agent is a nanomedicine.

22. A device for tumor priming with pressure pulses, the device comprising means for generating pressure pulses having a peak compressive pressure of 0.1 MPa to 100 MPa, with 90% of each pressure pulse lasting between 1 ns and 500 ns, for treating solid tumors in a subject suffering from cancer.

23. 23. A method according to claim 22, wherein the device is the device according to claim 1.

24. A system comprising at least one device according to claim 1.

25. 12. A kit comprising the device of any one of claims 1 to 11 and an anti-cancer therapeutic agent.

26. 26. The kit of claim 25, wherein the anti-cancer therapeutic agent is selected from the group consisting of an inhibitor of an inhibitory checkpoint molecule, an activator of a stimulatory checkpoint molecule, an antibody, a cytokine, an interferon, an interleukin, a vaccine, an oncolytic virus, a chimeric antigen receptor T cell, and any combination thereof.

27. 26. The kit of claim 25, wherein the therapeutic agent is a biological therapeutic agent.

28. 26. The kit of claim 25, wherein the therapeutic agent is a monoclonal antibody (mAb) used in cancer therapy, or any combination of mAbs used in cancer therapy.

29. 26. The kit of claim 25, wherein the therapeutic agent is selected from ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, cemiplimab, dostallimab, tislelizumab, leratolimab, toripalimab, camrelizumab, sintilimab, or any combination thereof.

30. 26. The kit of claim 25, wherein the therapeutic agent is a cytostatic or cytotoxic drug bound to a plasma protein.

31. 26. The kit of claim 25, wherein the therapeutic agent is a macromolecule.

32. 26. The kit of claim 25, wherein the therapeutic agent is a nanomedicine.

Images