Mirna composition accumulating in pancreas and / or spleen

Abstract

The disclosure provides a composition comprising one or more lipids and particles comprising hydrolysable silicon. The composition is for use in miRNA-based cancer treatment; in reducing or preventing accumulation of drug in the liver; in promoting accumulation of drug in the pancreas; and / or in promoting accumulation of drug in the spleen. Also provided are related compositions, compositions for use, and methods.

Description

[0001] MIRNA COMPOSITION ACCUMULATING IN PANCREAS AND / OR SPLEEN

[0002] Field

[0003] The present disclosure concerns delivery vehicles for active pharmaceutical ingredients (APIs; especially, miRNA). More particularly, but not exclusively, this disclosure concerns compositions comprising silicon particles, one or more lipids and an active pharmaceutical ingredient (API), especially, miRNA. The invention also concerns related products (including their uses) and methods.

[0004] Background

[0005] Micro RNA (miRNA) molecules are typically single stranded and non-coding. miRNA regulates a number of physiological processes, especially post-transcriptional regulation of gene expression, such as by mRNA silencing. In cancer, the abnormal downregulation of miRNA may result in pathological overexpression of oncogenes. miRNA replacement may therefore provide a therapeutic strategy. Additionally or alternatively, miRNA therapy may sensitise chemotherapy -resistant cancer cells to chemotherapeutic agents, enabling lower doses of such agents to be delivered, lowering their side effects in consequence. miRNA therapy may also be useful in treating other disorders.

[0006] However, a major difficulty in miRNA therapy lies in how to deliver miRNA in the correct amount and to the correct cells, a process known as targeted delivery.

[0007] Targeted delivery is important in the pharmaceutical field. It may provide specificity (reduced off-target effects); improve safety (reduced toxicity); and increase efficacy (by increasing the concentration of an API that reaches a target site), upon delivering an API to a patient in need thereof.

[0008] A common approach to targeted delivery has been to encapsulate an API into a lipid nanoparticle. Lipid nanoparticles may accumulate differentially in target tissues and thus distribute their API payload selectively to the target tissue. In some cases, it is thought that differential accumulation may be achieved by means of a so-called enhanced permeability and retention effect (known in the art as EPR). In EPR, high molecular weight API delivery platforms accumulate in any tissue offering increased vascular permeability, such as in sites of inflammation or cancer. However, EPR relies on a rudimentary, size-based effect that is not always predictable; nor is it particularly selective.

[0009] Additionally or alternatively to EPR, the outer surface of lipid nanoparticles may be decorated with one or more ligands with specificity for one or more targets, such as a particular cell having one or more complementary ligands.

[0010] Examples of such ligands include folic acid (binding to the folic acid receptor overexpressed in breast cancer and many other cancers); anisamide (which is thought to bind to the sigma receptor overexpressed by tumour-associated fibroblasts); tetraiodothyroacetic acid (capable of binding to the thyroid hormone receptor in renal cell carcinoma and breast cancer); and a ligand of the prostatic specific membrane antigen (for prostate cancer). Meanwhile, hyaluronic acid is used as a ligand with binding specificity for CD44 expressed by cancer stem cells. Other examples include antibodies (or fragments thereof) capable of binding to a variety of cancer- and cancer microenvironment-associated proteins (such, for example, as EGFR, HER2, PD-1, CD8, PD-L1 and VEGFR). Further examples include peptides, such as internalising RGD (iRGD; which binds to integrin and is internalized by neuropilin- 1, overexpressed by the endothelium of various cancers); or RGD (which simply binds to integrin). Aptamer ligands have also been developed, for example to target nucleolin or EPCAM (overexpressed in breast cancer).

[0011] The specificity of the ligand(s) leads to accumulation of the API payload near or within one or more specific tissues or even one or more specific cell types within a single tissue. However, this approach is not applicable to all cells (as the above examples show, much of the focus to date has been on certain types of cancer cells). Moreover, it requires the development of one or more new ligands for each new target.

[0012] At the same time, a fundamental issue with lipid nanoparticle-based API delivery vehicles lies in their potential for toxicity. This is because they typically require the presence of one or more exogenous lipids having a positive charge at physiological pH (known in the art as exotic cationic lipids). Such lipids may have an unfavourable toxicity profile, for example by triggering an immune response, especially when they are amine-rich. Although well-suited to the electrostatic loading of polyanionic nucleic acids (including various form of RNA) these (amine-rich) exotic lipid species may lead to cytotoxicity, immunogenicity, and may even lead, contrary to what is desired, to nonspecific tissue accumulation, thus counteracting the aim of targeted delivery. Furthermore, the types of lipids used in lipid nanoparticle-based API delivery vehicles tend to be complex to synthesise and are therefore expensive.

[0013] An additional drawback to lipid nanoparticle-based API delivery vehicles lies in their inability to stabilise fragile APIs. (As used herein, the terms “fragile APIs” and “reactive APIs” may be interchangeable and may refer to APIs which (i) have a halflife upon storage in aqueous solution at about 25 °C of at most one week, measurable by NMR or by GC-MS; and / or (ii) have a half-life in vivo of under about an hour, measurable by assay of a biological sample.) This may necessitate ultra cold chain storage of the lipid nanoparticle-encapsulated API, leading to an expensive and complex supply chain, especially when drug delivery to geographically remote territories or territories of an extreme climate is required.

[0014] Some organs, tissues and cells are especially difficult to target selectively.

[0015] One obstacle to organ, tissue and / or cell targeting in mammals, especially humans, is the liver. The liver is a major metabolic organ and plays a central role in detoxification of blood, in particular blood supplied by the hepatic portal vein. The liver may therefore be responsible for trapping and breaking down an API thus reducing the amount of API that reaches target cells or preventing the API from reaching target cells at all.

[0016] Meanwhile, an organ for which it is particularly difficult to trigger API accumulation naturally (i.e., in the absence of specific targeting strategies) is the spleen. The anatomy of the spleen is such as to present little or no sieving by endothelia. Thus, the amount of API that will enter into the spleen parenchyma is typically highly dependent on the amount of API capable of reaching the spleen via the blood. Although freshly ingested or injected API may in some instances reach the spleen before it reaches the liver, the API may quickly be metabolised by the liver, leading to a reduced steady state concentration developed over time in the blood and a reduced availability of the API to the spleen.

[0017] If a way could be found to target the spleen therapeutically, this would offer several advantages. In particular, the spleen acts as a gatekeeper in systemic immunity. For example, the role of the spleen in removing damaged and / or aged red blood cells makes the spleen conducive to therapeutic strategies involving erythrocyte decoration or “hitchhiking”. Monocyte precursors stored in the spleen may act as "Trojan horses" for shaping immune responses and / or delivering drugs to tumours (or other dysregulated organs, tissues or cells). Targeted delivery of APIs to the spleen therefore offers the potential for immunomodulation, involving the treatment of various immune- related conditions, including different cancer types, infectious diseases, and autoimmune disorders.

[0018] Another organ for which it is particularly difficult to trigger API accumulation naturally is the pancreas. Although the pancreas has an extensive blood supply (with vessels originating as branches of both the coeliac artery and superior mesenteric artery) and freshly ingested or injected API may in some instances reach the pancreas before it reaches the liver, the API may quickly be metabolised by the liver, leading to a reduced steady state concentration developed over time in the blood and a reduced availability of the API to the pancreas. Further hindrances include the heterogenicity of the pancreatic tissue; and its dense fibrotic stroma. Meanwhile, targeting the pancreas is desirable for the prevention or treatment of several widespread diseases and disorders, including cancer and diabetes.

[0019] There is therefore a need for improved accumulation of API in the spleen and / or pancreas.

[0020] There is also a need for new and improved ways to mitigate detrimental accumulation of an API in the liver, to maximise the API available to other organs (especially the spleen and / or pancreas). Returning to cancer, chemotherapy is still the only option for the treatment of many cancers (especially, metastatic pancreatic cancer). Chemotherapy also leads to significant and unpleasant side effects which in themselves may limit the amount of chemotherapy a patient is able or willing to undergo. It would be advantageous to develop a therapy capable of replacing or complementing chemotherapy, for example enabling tumour destruction to continue during pauses between chemotherapy treatment cycles, or during rest periods incorporated in chemotherapy treatment cycles, when the blood plasma concentration of the chemotherapeutic agent(s) is low or zero.

[0021] There is a need for new and improved means for treating cancer. There is a need for new and improved means for delivering miRNA to treat cancer, especially in a targeted way. There is a need for new and improved means for bypassing the liver to deliver miRNA to treat cancer.

[0022] The present invention seeks to mitigate the above-mentioned problems. Alternatively or additionally, the present invention seeks to provide an improved targeted delivery system for APIs (in particular, for RNA APIs, most especially miRNA).

[0023] Summary of the Disclosure

[0024] Provided herein is a composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of treating cancer by miRNA administration to a human patient in need thereof. The composition may further comprise (iii) the miRNA.

[0025] Also provided is a method of treating or preventing cancer, comprising administering to a human patient in need thereof a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) miRNA.

[0026] The present inventors have discovered that compositions comprising hydrolysable silicon with one or more lipids can deliver miRNA to adenocarcinoma cells in such a way as to reduce the formation and growth of tumour spheroids, i.e., inhibit tumour development, as evidenced by Examples 7a and 7b hereinbelow. This is surprising because not only is the miRNA delivered to the cells, but it is delivered to the correct location and intact to produce a therapeutic (anti-cancer) effect.

[0027] Meanwhile, Examples 3-5, 8 and 9 show the ability of these compositions to “bypass” the liver when carrying API, and reach other difficult-to-target organs, such as the spleen and pancreas. This is surprising because the liver commonly sequesters and metabolises API. It is concluded that these compositions would deliver miRNA to treat cancer cells in vivo while “bypassing” the liver, making more miRNA available to target organs, tissues and / or cells.

[0028] Thus, also provided is a composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of reducing or preventing accumulation of an active pharmaceutical ingredient (API) in the liver of a human patient. The composition may further comprise (iii) the API.

[0029] Especially, provided herein is a composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of treating cancer by miRNA administration to a human patient in need thereof, the method comprising reducing or preventing the accumulation of the miRNA in the liver.

[0030] Also provided herein is a method of reducing or preventing accumulation of an active pharmaceutical ingredient (API) in the liver of a human patient in need thereof, comprising administering to the patient a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) the API.

[0031] Especially, provided herein is a method of treating or preventing cancer, comprising administering to a human patient in need thereof a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) miRNA, the method comprising reducing or preventing the accumulation of the miRNA in the liver. As shown in Examples 3-5, 8 and 9, it is especially made possible to target difficult-to-target organs and, in particular, the pancreas and / or spleen.

[0032] Thus, provided herein is a composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of promoting accumulation of an active pharmaceutical ingredient (API) in the pancreas and / or spleen of a human patient in need thereof. The composition may further comprise (iii) the API.

[0033] Further, provided is a method of promoting accumulation of an active pharmaceutical ingredient (API) in the pancreas and / or spleen of a human patient in need thereof, comprising administering to the patient a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) the API.

[0034] It will of course be appreciated that features described in relation to one aspect of the present invention may be incorporated into other aspects of the present invention. For example, the method of the invention may incorporate any of the features described with reference to the apparatus of the invention and vice versa.

[0035] Description of the Drawings

[0036] In the accompanying drawings:

[0037] Figures la-b show IVIS imaging of anaesthetised albino CD1 mice in Example 3, at 6 hours and at 24 hours, respectively. The original colour images show a strong localised signal of bright green fluorescence in the pancreas and the spleen. Fluorescence of the liver is unexpectedly low (minimal to zero).

[0038] Figure 2 shows IVIS imaging of organs of culled albino CD1 mice in Example 3. Again, the original colour image shows bright green fluorescence, with a strong signal in the pancreas and also in the spleen. Fluorescence of the liver is unexpectedly low (again, minimal to zero). Figures 3a-l show whole-body live animal imaging and resected organ imaging carried out in Example 4. Bright red fluorescence is visible in the original colour images. In whole body imaging, the signal from the intraperitoneal site of injection is strong (and may mask signals from other locations). In the resected organs, the spleen signal is unexpectedly strong, while the liver signal is unexpectedly weak.

[0039] Figure 4 shows siRNA quantification in resected organs over time (measured using SL-qPCR) in Example 4. Again, the spleen signal is strong, while the liver signal is weak. The results shown in Figure 4 are not normalised by organ weight, hence the outsize signal for the kidney; and the localisation to the (small) spleen may be even more pronounced, compared to the (larger) liver signal, than the unnormalized data shows.

[0040] Figures 5a-c show bioluminescence results for Group 4 mice in Example 5.

[0041] Figures 6a-c show fluorescence results for Group 5 mice in Example 5. The original colour images show bright red fluorescence. The signal for the spleen and pancreas was unexpectedly strong, while that for the liver was unexpectedly weak.

[0042] Figures 7a-c show fluorescence results for Group 6 mice in Example 5. Again, the original colour images show bright red fluorescence. The signal for the spleen and pancreas was strong, while that for the liver was weak.

[0043] Figures 8a-c show fluorescence results for Group 7 mice in Example 5. Again, the original colour images show bright red fluorescence. The signal for the spleen and pancreas was strong, while that for the liver was weak.

[0044] Figures 9a-c show bioluminescence results for Group 8 mice in Example 5. The original colour images show bioluminescence in a scale from blue (low) through green (medium) to high (red); against a white-grey background of no bioluminescence. The trend in bioluminescence suggests preferential spleen and pancreas uptake. Some bioluminescence is observed in the tail (site of injection). Figures lOa-c show bioluminescence results for Group 9 mice in Example 5. Again, the original colour images show bioluminescence in a scale from blue (low) through green (medium) to high (red); against a white-grey background of no bioluminescence, with preferential spleen and pancreas uptake.

[0045] Figures lla-c show bioluminescence results for Group 10 mice in Example 5. Again, the original colour images show bioluminescence in a scale from blue (low) through green (medium) to high (red); against a white-grey background of no bioluminescence, with preferential spleen and pancreas uptake.

[0046] Figure 12 compares graphically the luminescence observed for the liver vs. the spleen in Example 6. Comparison of the spleen to liver luciferase activity revealed statistically significant targeting of the spleen over the liver for at least test groups 4, 5 and 7, as shown in Figure 12. There appears to be a similar trend for group 6, but it is somewhat less significant. Representative results are shown in further detail in Figures 13a-c.

[0047] Figures 14 to 16 show the results of miRNA delivery to cancer cells, using compositions in the scope of the present disclosure, over time. Successful delivery of miRNA to the cells was evidenced by inhibition of tumour development by the miRNA.

[0048] Figure 17 is a transmission electron microscope (TEM) image that shows the aggregation of silicon particles, as described herein.

[0049] Figure 18 shows fluorescence results for mice in Example 8. The original colour images show bioluminescence in a scale from blue (low) through green (medium) to high (red); against a white-grey background of no bioluminescence. The trend in bioluminescence suggests preferential spleen and pancreas uptake, with a surprisingly minimal or zero liver uptake. Some bioluminescence is observed in the tail (site of injection).

[0050] Figures 19a-c and 20 show the luminescence and fluorescence results for mouse organs in Example 9. Detailed Description

[0051] While the subject-matter of the present disclosure is described and illustrated below with reference to particular embodiments, it will be appreciated by those skilled in the art that the subject-matter lends itself to many different variations not specifically illustrated herein. By way of example only, certain possible variations will now be described.

[0052] Particles comprising hydrolysable silicon

[0053] The composition disclosed herein comprises particles comprising or consisting of hydrolysable silicon.

[0054] At least about half of the silicon atoms in the particles may be in the form of elemental silicon, which is hydrolysable.

[0055] Preferably, the particles comprising hydrolysable silicon contain at least about 50 % by weight silicon atoms relative to the total weight of the particles. They may contain at least about 60, at least about 70, at least about 80, at least about 90, and especially at least about 95 % by weight silicon atoms relative to the total weight of the particles comprising the hydrolysable silicon.

[0056] The particles comprising hydrolysable silicon may thus be formed of pure or substantially pure silicon present as the element. Preferably, the particles have minimal surface oxidation at most; and have no oxidation in their cores.

[0057] Although the particles may contain traces of silica, silica is not hydrolysable silicon. Any given sample of silica contains less than 47 % by weight silicon (the molecular mass of silicon is 28 and the molecular mass of oxygen is 16; 100 x 28 / (16+16+28) = 46.67).

[0058] A sample of the hydrolysable silicon may show a rate of hydrolysis, for example in PBS buffer at room temperature, of at least about 10% (preferably, at least about 50 %) of the rate of hydrolysis of a sample of pure silicon (the element) of the same dimensions. Assays for hydrolysis of silicon-containing material are widely known in the art; see, for example, WO 2011 / 001456, incorporated herein by reference in its entirety.

[0059] The particles may be surface treated with one or more alcohols, such as methanol, benzyl alcohol or ethanol. This may be or comprise the formation of Si-OC- containing groups, such as Si-O(CH2)XCH3 (it will be understood that x is zero for methanol, 1 for ethanol, 2 for propanol, etc.; and (CH2)xmay be replaced by other carbon-containing groups, such as an aromatic ring, for example when the alcohol is or comprises benzyl alcohol), Si-(CH2)XCH3 and / or Si-H-containing moieties on the particles’ surface.

[0060] The silicon-containing compositions disclosed herein for delivery of an API (especially, an API which is miRNA) are preferably non-toxic. For example, they may be substantially fully biodegradable as described herein, in that the silicon may degrade to non-toxic orthosilicic acid in vivo.

[0061] The silicon-containing compositions disclosed herein for delivery of an API (especially, an API which is miRNA) may be fully dispersible in an aqueous environment, to ensure ease of delivery, such as by injection in aqueous solution. Accordingly, the compositions may further comprise an aqueous carrier, such as water. Also provided herein is an aqueous dispersion, comprising the silicon-containing compositions disclosed herein. Optionally, the aqueous dispersion is formulated for injection or ingestion, or for topical application, by or to a human patient in need thereof.

[0062] Additionally or alternatively, the composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, as it is disclosed herein, may be dispersed in a gel (such as a hydrogel and / or a biodegradable gel). It may especially be dispersed in a gel comprising sodium hyaluronate or a derivative thereof. Optionally, the gel is formulated for injection or ingestion, or for topical application, by or to a human patient in need thereof. Shape and surface characteristics of the particles

[0063] The particles may, especially, be nanoparticles. The nanoparticles may have a nominal diameter of about 1 to about 500 nm, especially about 1 to about 250 nm, more especially about 1 to about 100 nm (e.g., about 10 nm, 30 nm or 50 nm). As used herein, the term “nominal diameter” may refer to the mean diameter and at least about 90 % of total mass of particles in a sample of particles may fall within the size range specified.

[0064] The particles may be porous, especially mesoporous. As used herein and in accordance with IUPAC nomenclature, the term mesoporous may mean that the particles are nanoparticles (with a mean diameter in a range of from about 1 to about 500 nm) having pores with diameters in a range of from about 1 to about 50 nm.

[0065] The particles’ size and porosity may be measured, for example, by scanning electron microscopy.

[0066] Particles comprising hydrolysable silicon can be made porous by standard techniques such as contacting the particles with a hydrofluoric acid (HF) / ethanol mixture and applying a current. By varying the HF concentration and the current density and time of exposure, the density of pores and their size can be controlled and can be monitored, such as by scanning electron microscopy. If the particles are porous, their total surface area will be increased by virtue of their porosity. For example, their surface area may be increased by at least about 50 % or at least about 100 %, compared to the surface area of a non-porous but otherwise identical particle. In many circumstances, porous particles will in reality have a much greater increase in total surface area by virtue of their porosity. According to certain embodiments the porosity is at least about 30, about 40, about 50 or about 60 %; meaning that, respectively, at least about 30, about 40, about 50 or about 60 % of the particle volume is pore space. Pore diameters may be in a range of from about 1 nm to about 50 nm, for example from about 1 nm to about 5 nm.

[0067] It will be appreciated that the particles may be produced by various techniques familiar to the skilled person. The techniques may include, for example, purely physical (sometimes referred to, in the art, as “non-we ’) processes having bulk silicon (especially, silicon wafer) as the starting material; such as pulsed laser ablation, thermal degradation and ball milling. Thus, the particles may be obtainable by a method comprising or consisting of one or more of pulsed laser ablation, thermal degradation and ball milling, of bulk silicon (especially, silicon wafer).

[0068] Additionally or alternatively, the particles may be produced by chemical (sometimes referred to, in the art, as “wet”) techniques, including but not limited to electrochemical etching of bulk silicon (especially, silicon wafer). Such techniques optionally include the HF etching described above. Thus, the particles may be obtainable by a method comprising or consisting of electrochemical etching of bulk silicon (especially, silicon wafer).

[0069] Once formed, silicon particles may be sorted by size, such, for example, as by air classification, sieving and / or filtration. Thus, the particles may be obtainable by a method comprising one or more of air classification, sieving and / or filtration.

[0070] Thus, for example, the particles may be obtainable by a method comprising producing silicon particles from bulk silicon, especially from silicon wafer, such as by one or more of pulsed laser ablation, thermal degradation, ball milling, and electrochemical etching, of bulk silicon (especially, silicon wafer); followed, optionally, by sorting by size, such as by air classification, sieving and / or filtration.

[0071] Optionally, the particles may be washed before use, such as in methanol or ethanol, to remove a thin oxidised layer from their surface. In the art, this may be termed “activation”.

[0072] The particles thus obtained may have a narrow size distribution and homogeneous surface chemistry, leading to batch-to-batch reliability and reproducibility of one or more of the advantages described herein. Suitable physical and chemical techniques are set out, for example, in WO 2011 / 012867 Al (in the name of SISAF LTD); in Tokarska K et al., Facile production of ultra-fine silicon nanoparticles, R. Soc. Open Sci., 2020, 7: 200736; and in Kim, T., Lee, J. Silicon nanoparticles: fabrication, characterization, application and perspectives, Micro and Nano Syst. Lett, 2023, 11: 18, each of which is incorporated herein by reference in its entirety.

[0073] Doping of the silicon

[0074] The particles may comprise hydrolysable doped silicon. As used herein, the term “doped silicon” may refer to silicon which behaves as an extrinsic semiconductor due to the presence of dopant atoms. The dopant atoms may be or comprise dopant atoms which are substitutional (taking the place of Si atoms). Additionally or alternatively, the dopant atoms may be or comprise dopant atoms which are interstitial (amongst, not displacing, Si atoms). Optionally, the dopant atoms may be present on the surface of the particles. Optionally, the dopant atoms may be present only on, or only on or close to, the surface of the particles. In this way, the particle may optionally have an undoped silicon core, while having an outer shell, or a surface, which is doped.

[0075] However, preferably, no metal silicate coating is present on the surface of the particles. A surface coating of metal silicate is not doping. Unlike scattering dopant atoms amongst silicon atoms, a surface coating of metal silicate would cover the surface of the particle with the metal silicate compound. Although silicon particles can be coated with metal silicate, to do so may complicate their synthesis and is not necessary for the beneficial effects described herein, such as but not limited to tissue targeting.

[0076] The particles may be doped at a level of at least about 1 xlO15, most especially at least about 1 xlO16dopant atoms per cm3. For example, the particles may be doped at a level of at least about 1 xlO17, at least about 1 xlO18, or at least about 1 xlO19dopant atoms per cm3. The particles may be doped at a level of up to 1 xlO20dopant atoms per 3 cm .

[0077] The silicon particles may be n-doped or p-doped. The silicon particles may be doped with one or more elements selected from B, P, Mg, Cu, Ga, Al, In, Bi, Ge, Li, Xe, N, Au and Pt. Thus, the dopant may be a p-dopant, especially boron. The dopant may be an n-dopant, especially phosphorus.

[0078] The dopant may be incorporated in a hydrolysable silicon matrix, (i) substitutionally, (ii) interstitially and / or (iii) via surface attachment. In this way, an improved API delivery vehicle may be provided.

[0079] When boron is used as the dopant, as is preferred, doping levels of IxlO15dopant atoms per cm3, and IxlO20dopant atoms per cm3may correspond, respectively, to a resistivity of 13.6 Q-cm, and 1.3 m Q-cm. Embodiments wherein boron is the dopant do not exclude silicon which, while doped (e.g., heavily doped) with boron, is additionally doped with other elements (preferably, in such cases, the majority dopant is boron).

[0080] As used herein, the term “heavy doping” is understood to mean doping of at least about 1 xlO15dopant atoms per cm3. In some preferred embodiments, dopant is present at levels of at least about 1 xlO16dopant atoms per cm3. Thus, in particularly preferred embodiments, the dopant is boron which is present at levels of at least about IxlO16boron atoms per cm3. For example, there may be boron present at levels of at least about 1 xlO16boron atoms per cm3and up to about 1 xlO20boron atoms per cm3.

[0081] Where silicon is referred to herein as “undoped” (such as the particles of compositions 2C-SiAEN-RE0.1 or 3C-SiAEN-RE0.1 of the Examples below), it may mean that no or only small amounts of dopant atoms are present, for example at most about IxlO2dopant atoms per cm3. Additionally or alternatively, “undoped” silicon may mean silicon that does not behave as an extrinsic semiconductor.

[0082] Advantageously, doping the particles comprising hydrolysable silicon may have a beneficial effect on the zeta potential of the particles, which may be a proxy for the surface charge of the particles. Doping may provide a zeta potential more suitable for improved loading of an API. It is thought that this may help stabilise an API while it is in circulation in vivo until it reaches a target cell, where it is subsequently released. Thus, more API may reach a target cell in a given time period after administration, compared to when undoped particles are used, leading to more efficient API delivery.

[0083] Doping the particles may increase the half-life in vivo (at about pH 7.4 and about 37 °C) of the API, such as by a factor of about 1.5, especially a factor of about 2. Especially, in the presence of the particles comprising hydrolysable doped silicon and the one or more lipids, the API may have a half-life in vivo of more than about 1 hour, especially more than about 2 hours, more especially more than about 6 hours. It will be appreciated that the term “half-life in vivo of the API”, as used herein, may refer to the elimination half-life in vivo of the API, i.e. the time period taken for the amount of the API, once administered, to reduce by about half. It will also be appreciated that the term “amount of the API” may refer to the amount of the API or a derivative thereof having the same or substantially the same intended pharmaceutical effect. It will also be appreciated that the term “half-life in vivo", as used herein, may refer to a systemic half-life of the API (inside the body, especially the human body); as opposed to the half-life of the API on an external body surface, such as when present on the skin surface (outside the body, even if in contact with it).

[0084] The doping of the particles may optionally change their zeta potential by a magnitude of at least about 5 mV, especially at least about 10 mV, compared to undoped particles.

[0085] Modulation of the zeta potential of the particles (by doping; and / or by the presence of the one or more lipids) may assist the particles to bypass the liver; to avoid the detrimental accumulation of the API in the liver; and / or to promote accumulation of the API in the pancreas and / or spleen. Thus, it may help with targeting of the API to an organ, tissue or cell of interest.

[0086] Doping with a p-dopant may lead to a more positive zeta potential, leading to improved binding with an API having a net negative charge, especially an API having a net negative charge at a pH of about 7.4 (because this is a typical physiological pH), especially a nucleic acid, more especially mRNA, siRNA or miRNA, most especially miRNA. Thus, in some embodiments, the hydrolysable doped silicon particles are doped with a p-dopant and the API is a negatively charged API, especially a nucleic acid, more especially mRNA or miRNA, most especially miRNA.

[0087] P-doping may still provide improved binding with an API having a net positive charge, especially an API having a net positive charge at a pH of about 7.4 (because this is a typical physiological pH), especially where the particles’ zeta potential is further modulated by other components such as the one or more lipids (e.g., phospholipid(s)) being present. Without wishing to be bound by theory, it is thought that electron transfer may occur from p-doped silicon (e.g., boron-doped silicon) to other constituents, thereby modulating particle size and charge, and hence nucleic acid binding capability.

[0088] The p-doping of the particles may optionally increase their zeta potential by at least about 5 mV, especially at least about 10 mV, compared to undoped particles.

[0089] Meanwhile, doping with an n-dopant may lead to a more negative zeta potential, leading to improved binding with a positively charged API. Thus, in some embodiments, the particles comprising hydrolysable doped silicon are doped with an n- dopant and the API is an API having a net positive charge, especially an API having a net positive charge at a pH of about 7.4.

[0090] N-doping may still provide improved binding with an API having a net negative charge, especially an API having a net negative charge at a pH of about 7.4 (because this is a typical physiological pH), especially where the particles’ zeta potential is further modulated by other components such as the one or more lipids (e.g., cationic lipid(s)) being present. Additionally or alternatively, n-doped silicon may be able to protect the one or more lipids, especially zwitterionic or positively charged lipids, from degradation. This may indirectly help to bind and stabilise an API, for example a nucleic acid, even where the API is negatively charged (especially, mRNA or miRNA, most especially miRNA).

[0091] The n-doping of the particles may optionally decrease their zeta potential by at least about 5 mV, especially at least about 10 mV, compared to undoped particles. P-doping or n-doping of particles comprising hydrolysable silicon may lead to improved binding with zwitterionic or neutral APIs. Thus, in some embodiments, the particles comprise hydrolysable p-doped silicon and the API is zwitterionic or neutral. In other embodiments, the particles comprise hydrolysable n-doped silicon and the API is zwitterionic or neutral.

[0092] Conventional liposomal transfection compositions (which do not comprise silicon) tend to comprise a significant quantity (such as a majority % by weight) of cationic lipid, such as DOTMA, wherein the positive charge of the cationic lipid is intended to stabilise negatively charged APIs, particularly nucleic acid. However, cationic lipid may not be cost-effective and may potentially not have an adequate safety profile for all clinical applications, such as administration to certain patient groups, e.g. infants, the elderly, or pregnant women. As described herein, doped (especially, p- doped) silicon provides the potential to use less or no cationic lipid, thus may provide improved cost-effectiveness and safety, e.g. enabling administration to infants, the elderly, or pregnant women.

[0093] Doping of the particles may help maintain a more positive or more negative (as the case may be) zeta potential than otherwise, meaning that components having certain functionalities can be added that would otherwise increase / decrease (as the case may be) the particles’ zeta potential too much, to maintain satisfactory API binding and delivery.

[0094] The manufacture of doped silicon is well-understood in the semiconductor industry and includes ion implantation and diffusion methods, making doped silicon per se readily available. As an example of a diffusion method, silicon powder and a doping reagent (for example B2O3 for boron doping) are mixed under N2 atmosphere at a temperature of 1050 °C - 1175 °C for a few minutes, to allow the dopant (such, for example, as boron) to diffuse into the silicon. Lipids

[0095] The one or more lipids may be or comprise one or more (preferably all) of one or more cationic lipids; one or more phospholipids; and one or more polyethylene glycol lipids (PEG lipids, or PEGylated lipids; the terms are used interchangeably).

[0096] As used herein, the term “cationic lipid” may refer to a lipid having a positive charge at physiological pH (especially, in vivo in humans). Examples include ditetradecyl trimethyl ammonium (DTDTMA), 2,3-dioleyloxypropyl-l-trimentyl ammonium (DOTMA), dihexadecyl trimethyl ammonium (DHDTMA); dioleoyl-3- trimethylammonium propane (DOTAP); DODAP (l,2-dioleoyl-3-dimethylammonium propane) and stearylamine (SA). Preferred examples are DODAP and DOTAP; most preferred is DOTAP. DOTAP exists in an S and an R enantiomeric form, and may be present as substantially the S- form, present as substantially the R- form, or present as a racemate.

[0097] The term “phospholipid” may refer to a lipid containing at least one phosphate group. The term encompasses glycerophospholipids. The term encompasses mixtures of phospholipids, including mixtures of phospholipids found in lecithin. The phospholipid may contain one or more aliphatic side chain(s) and / or one or more ether side chains (ether side chains include polyethylene glycol chains and polypropylene glycol chains). Examples of phospholipids include phosphatidylcholine (PC), hydrogenated PC, dioleoylphosphatidylethanolamine (DOPE) and dipalmitoylphosphatidylcholine (DPPC). A preferred phospholipid is DOPE. A phospholipid may have a net neutral charge, such for example as a zwitterionic phospholipid. A phospholipid may have a net negative charge. Optionally, the phospholipid(s) is (or are) non-cationic. In the art, phospholipids are typically classified as neutral lipids.

[0098] The term “PEG lipid” may refer to a PEG-lipid conjugate in which polyethylene glycol is covalently bonded to a lipid molecule, such as l,2-distearoyl-sn-glycero-3- phosphoethanolamine (DSPE); a preferred example is DSPE-PEG2000.

[0099] The one or more lipids may be selected from one or more of: di oleoyl-3 -trimethylammonium propane (DOTAP) l,2-dioleoyl-3 -dimethylammonium propane (DODAP)

[0100] 9-heptadecanyl 8-{(2-hydroxyethyl)[6-oxo-6-

[0101] (undecy 1 oxy )hexy 1 ] amino } octanoate ( SM- 102) dioleoylphosphatidylethanolamine (DOPE)

[0102] PEGylated l,2-distearoyl-sn-glycero-3-phosphoethanolamine (e.g.

[0103] DSPE-PEG2000)

[0104] PEGylated diethylenetriamine (e.g. DTA-PEG2000) and derivatives thereof.

[0105] Optionally, the one or more cationic lipids encompass lipids which are only positively charged when they are protonated, such as the ionisable lipid SM-102 (9- heptadecanyl 8-{(2-hydroxyethyl)[6-oxo-6-(undecyloxy)hexyl]amino}octanoate) which is neutral unless its amine group is protonated.

[0106] The one or more lipids may preferably comprise or consist essentially of a combination of cationic lipid and phospholipid, such as the combination of DOTAP and DOPE. Optionally, the one or more lipids comprise or consist essentially of a mixture of DOTAP, SM-102 and DOPE.

[0107] The one or more lipids may optionally comprise or consist essentially of a combination of DOTAP, DOPE and a PEG-lipid (especially DSPE-PEG2000).

[0108] Optionally DODAP may be present in addition to or instead of DOTAP.

[0109] When the one or more lipids comprise or consist essentially of a combination of cationic lipid with phospholipid, especially the combination of DOTAP and DOPE, or the combination of DOTAP, DODAP and DOPE, the ratio by weight of cationic lipid: phospholipid (e g., DOTAP : DOPE, or DOTAP + DODAP : DOPE) may be in a range of from about 1 :3 to about 3: 1; such, for example, as about 1 :2 to about 2: 1. Examples of compositions having such a ratio include 3C-SiAEN-RE0.1, 3C-SiLB- RE0.1, 2C-SiAEN-RE0.1, 2C-SiAEN-Tyr-RE0.1, 2C-SiLB-RE0.1, 2C-LBSi-Tyr- RE0.1, 2C-LBSi-Tyr-RE0.1 tagged with DOPE-CF, 2C-AESi-Tyr-RE0.1, 2C-LBSi- BPEHO-Tyr-REO. l with 0.5% trehalose, 3ISC30.70-P159.3-AESi-SA-RE0.1, 3ISC30.70-P159.3-LBSi-SA-RE0.1, 2C-LBSi-Ch-OSA0.3-Arg-Gly-SA-RE0.1, 2C- LBSi-Ch-OSA0.3-Tyr-Gly-RE0.1 and 2C-AESi-TFF0.1 of the Examples below.

[0110] When the one or more lipids comprise or consist essentially of a combination of cationic lipid (e.g. DOTAP) with PEGylated lipid, the ratio by weight of cationic lipid: PEGylated lipid (especially, DOTAP: PEGylated lipid) may be in a range of from about 50: 1 to about 2: 1; such, for example, as about 10: 1 to about 2: 1. Examples of compositions having such a ratio include 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1 of the Examples below.

[0111] When the one or more lipids comprise or consist essentially of a combination of phospholipid (e.g., DOPE) with a PEGylated lipid, the ratio by weight of phospholipid: PEGylated lipid (e.g., DOPE: PEGylated lipid) may be in a range of from about 50: 1 to about 2: 1; such, for example, as about 10: 1 to about 2: 1. Examples of compositions having such a ratio include 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1 of the Examples below.

[0112] The one or more lipids may be bound (especially non-covalently) to the surface of the particles. The one or more lipids may be complexed with the API, such as a nucleic acid (especially, mRNA, siRNA or miRNA, most especially miRNA).

[0113] Preferably, the one or more lipids is or are substantially non-toxic. I.e., the one or more lipids preferably exclude toxic lipids. Preferably, the one or more lipids substantially do not stimulate the innate immune system in humans. Such lipids are known and include various phospholipids, non-toxic cationic lipids, and various PEG lipids; such as DOPE, DOTAP and DSPE-PEG2000

[0114] The one or more lipids may exclude exogenous lipids having a large (e.g., 3+, 4+ or above) net positive charge at physiological pH (known in the art as exotic cationic lipids) and / or which are amine-rich; such lipids may have an unfavourable toxicity profile, for example by triggering an innate immune response upon administration to a human patient, especially when they are amine-rich. Although well-suited to the electrostatic loading of polyanionic nucleic acids (including various form of RNA) these (amine-rich) exotic lipid species may lead to cytotoxicity, immunogenicity, and may even lead, contrary’ to what is desired, to non-specific tissue accumulation, thus counteracting the aim of targeted delivery. Furthermore, they may be complex to synthesise and may therefore be expensive.

[0115] Thus, the one or more lipids may exclude amine-rich lipid. It will be appreciated that the DOPE, DODAP, DOTAP, SM-102 and DSPE-PEG2000 of the Examples below are not amine-rich in this sense. Amine-rich lipids may be defined as having more than 2, 3 or 4 nitrogen atoms per molecule of lipid. In contrast, the one or more lipids in accordance with the present disclosure preferably contain up to 1 nitrogen atom per molecule of lipid.

[0116] It has been found that the beneficial effects disclosed herein may be achieved without necessarily being tied to one or more specific lipid compounds. The one or more lipids may play a role in charge-charge interactions, such as in modulating the zeta potential at the surface of silicon particles, so as to enable better binding of the API. This effect may be seen across a wide range of lipids. Lipids as a class show trends in properties, especially in terms of inter-molecular interactions, enabling the compositions of the present disclosure to be implemented with different lipids and in differing amounts, compared to the specific lipids disclosed in the Examples below. In particular, DOPE is a representative example of the class of phospholipids, to which its behaviour may be extrapolated. DOTAP is a representative example of the class of cationic lipids, which is also modelled to some extent by SM-102 (although SM-102 is neutral until protonated, upon protonation it behaves as a cationic lipid). DSPE-PEG2000 is a representative example of the class of polyethylene glycol (PEG) lipids.

[0117] Actually, the compositions of the present disclosure may enable less cationic lipid, such as DOTAP and / or DODAP, to be used, compared to conventional compositions for API delivery (such as lipid nanoparticles which comprise cationic lipid). While cationic lipids may be suitable for electrostatic loading of polyanionic nucleic acids (including various form of RNA) their amine-rich nature may cause cytotoxicity, immunogenicity, and non-specific tissue accumulation. Thus, the one or more lipids may optionally exclude cationic lipid.

[0118] Thus, the one or more lipids may be or comprise one or more of: one or more phospholipids (e.g. DOPE); and one or more polyethylene glycol (PEG) lipids (e.g. DSPE-PEG2000).

[0119] Overall, the hydrolysable silicon-containing particles disclosed herein can provide the potential to use less lipid (especially less cationic lipid, such as less DOTAP) in API delivery vehicles, compared to conventional API delivery vehicles which do not contain hydrolysable silicon particles (e.g. conventional liposomal nucleic acid delivery vehicles, such as those typically used for mRNA delivery in vivo). Additionally or alternatively, the hydrolysable silicon particles can provide the potential for API delivery vehicles to be formulated with a wider range of lipids while still providing transfection efficiency, storage stability, and / or targeted delivery to a particular type of tissue, or to a particular type of cell. In turn, this may lead to reduced reliance in the field on specific lipids, particularly cationic lipids, especially cationic lipids which are formulated specifically for the purpose of API delivery and which may therefore not be cost-effective or easily accessible.

[0120] It has been found that the one or more lipids may help to control the rate of release of the API (e.g. nucleic acid, especially mRNA, miRNA or siRNA, most especially miRNA). The type of lipid used may help modulate the rate of API release. Thus, different lipids may be used in different amounts to achieve a given rate of API release.

[0121] The one or more lipids may have a beneficial effect on the surface charge of the particles. They may provide a zeta potential more suitable for improved loading of an API, especially a nucleic acid (such, for example, as miRNA, siRNA, short activating RNA, short hairpin RNA or mRNA, most especially miRNA). They may help to control the rate of API release at a target site. Thus, different lipids may be used in different amounts to achieve a given zeta potential. By way of example, the addition of DOTAP to particles comprising hydrolysable pure silicon may promote a more positive surface charge, thus modulating the particles’ zeta potential.

[0122] The one or more lipids may help to modulate the rate of hydrolysis of the silicon, such that the silicon hydrolyses to bioavailable orthosilicic acid (OSA) degradation product; rather than insoluble polymeric hydrolysis products. Controlling the rate of hydrolysis of the silicon may influence the rate of release of API associated with the silicon. Controlling the rate of API release may modulate the length of the time period during which protection of the API is sustained, especially concerning protection in vivo in the presence of various bodily fluids. Thus, more API may be delivered to a target cell in a given time period, than for an otherwise identical composition.

[0123] Without wishing to be bound by theory, while some lipids are investigated in the Examples hereinbelow, the mechanism by which the one or more lipids act may be due to properties of the class of lipids, such as their behaviour in response to chargecharge interactions; and may thus be generalisable beyond the exemplified lipids. Thus, different lipids may be used in different amounts to achieve a given zeta potential and / or a given rate of API release and / or a given rate of hydrolysis of the silicon.

[0124] Lipids are generally understood to include fatty acids and fatty acid derivatives, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids and polyketides. As used in the present application, the term “lipid” may nevertheless encompass lipidated oligopeptide (a term used interchangeably herein with the term lipopeptide) wherein a short peptide sequence (such as a peptide sequence having 3 to 20 amino acid residues, such as 5 to 15 amino acid residues, especially 3, 4, or 5 amino acid residues, and most especially 5 amino acid residues) is conjugated to one or more fatty acid chains (especially a fatty acid chain having a 10 to 24 carbon chain length, preferably, a 12 to 18 carbon chain length; for example a 14, 15 or 16 carbon chain length; for instance, the peptide moiety may optionally be lipidated with a palmitoyl, cetyl or myristoyl moiety). The one or more lipids may thus comprise one or more lipidated oligopeptides. Preferably, the one or more lipidated oligopeptides each comprise a fatty acid chain having in the range of about 12 to about 18 carbon atoms.

[0125] Preferably, the one or more lipidated oligopeptides each comprise 3 to 20 amino acid residues. Thus, the lipidated oligopeptide may be a lipidated tetrapeptide, lipidated pentapeptide or lipidated hexapeptide.

[0126] Preferably, the amino acid residues include at least one amino acid residue (for example, about 2 or about 3 amino acid residues) that is cationic at a pH of about 7.4 (physiological pH), such, for example, as lysine or arginine. For example, the lipidated oligopeptide may include one or more (for example, about 2) lysine resides.

[0127] An especial example of a lipidated oligopeptide (“lipopeptide”) is palmitoyl- pentapeptide-4 (CAS number 214047-00-4; abbreviated as PAL-KTTKS).

[0128] Thus, the one or more lipids may comprise or be one or more lipidated oligopeptides, particularly those having one or more amino acid residues that is or are positively charged at a pH of about 7.4 (i.e., about physiological pH) such, for example, as one or both of lysine and arginine.

[0129] The lipidated oligopeptide may especially be used in combination with one or more phospholipids, such as DOPE. The alkyl chain of a lipidated oligopeptide molecule may be assimilated in a phospholipid bilayer, while the surface of the bilayer is decorated with the peptide moiety. Without wishing to be bound by theory, it is thought that a peptide moiety of the lipidated oligopeptide can enhance the targeting of one or more specific tissues and / or cells. Meanwhile, where the peptide moiety bears a positive charge at a pH of about 7.4 (i.e., about physiological pH), it may stabilise negatively charged APIs (e.g. nucleic acids, especially mRNA, siRNA or miRNA).

[0130] The one or more lipids may be or comprise one or more structural lipids (e.g. a cholesterol-based lipid). However, the one or more lipids may optionally exclude structural lipid. Thus, the one or more lipids may exclude sterols; especially, they may exclude cholesterol. It has been found that the presently disclosed compositions need not rely on these types of lipid, which traditional API delivery systems typically rely on. Thus, the compositions disclosed herein have the potential to provide alternatives to API delivery systems reliant on these types of lipids, especially cholesterol. Where cholesterol is not available or where its use is otherwise not possible (e.g., due to its effect in the body) this may be advantageous.

[0131] The one or more lipids may have an average molecular weight of about 100 to about 2000, such as about 500 to about 1000.

[0132] The ratio by weight of the total lipids (by which is meant all lipid components in the composition, taken together) to silicon may be in a range of from about 50:1 to about 1 : 1, especially a range of about 30 : 1 to about 1 : 1; such, for example, as a ratio by weight of about 16: 1. This may suitably be determined at a point when the components are assembled for manufacture of a delivery system, i.e. before any further processing is carried out (such further processing may be, for example, the processing under “Extrusion” of the Examples hereinbelow).

[0133] Configurations that may be adopted by compositions described herein

[0134] The composition of the present disclosure preferably comprises or is formed of particulates. The particulates may have a diameter in a range of about 10 nm to 1000 nm, such as about 50 to about 500 nm. The particulates may comprise at least one hydrolysable silicon particle (or an aggregation of hydrolysable silicon particles) for example at least about 5, 10, 20 or 50 hydrolysable silicon particles, bound together with one or more other components disclosed herein - especially, bound together with the one or more lipids and / or the API (most especially miRNA).

[0135] Preferably, the particles comprising hydrolysable silicon are bound non- covalently to the one or more lipids, thus forming a delivery vehicle for transport of the API. In turn, preferably, the API is bound non-covalently to the particles comprising hydrolysable silicon. When other components (especially, amino acid(s)) is or are present, they may form a complex together with the particles, lipid(s) and / or API. Thus, the composition may be in the form of a complex of the particles, lipid(s), amino acid(s) and API.

[0136] The one or more lipids may be formed of, or may comprise, one or more lipid structures. Said structures may be or comprise one or more of: micelles, incomplete micelles, liposomes, incomplete liposomes and (e.g., semi-solid or substantially solid) lipid globules, as described herein.

[0137] Meanwhile, preferably, the API is bound (especially, bound non-covalently) to the particles comprising hydrolysable silicon. Preferably, at least about 50, 60 or 70 % of API in the composition is bound to the particles in this way.

[0138] In turn, preferably, the particles comprising hydrolysable silicon are bound to the surface of, and / or are present in the interior of, the one or more lipid structures.

[0139] Thus, preferably, particles comprising hydrolysable silicon are bound to the surface of, and / or are present in the interior of, one or more of: micelles, incomplete micelles, liposomes, incomplete liposomes, and (e.g., semi-solid or substantially solid) lipid globules; and API is bound to the particles comprising hydrolysable silicon. Preferably, the particles comprising hydrolysable silicon are present in one or more aggregates of particles comprising hydrolysable silicon, especially one or more aggregates comprising or consisting of chains of the particles, most especially chains that extend into the interior of the one or more lipid structures, such as into the interior of one or more of: micelles, incomplete micelles, liposomes, incomplete liposomes, and (e.g., semi-solid or substantially solid) lipid globules (especially, liposomes and / or lipid globules). An amino acid (especially, glycine, arginine and / or tyrosine, such as glycine) may also be associated with (e.g., bound non-covalently to) the particles comprising hydrolysable silicon; and / or associated with (e.g., on the surface of) the one or more lipid structures.

[0140] As used herein, the term liposomal lipid particle, or the term liposome, may have its normal meaning in the art. Thus, it may refer to a vesicle having at least one lipid bilayer, which may be approximately spherical in shape. A liposome may be visualised as a lipid “bubble” surrounding an interior space. The interior space may be a hydrophilic environment.

[0141] The composition may in some embodiments comprise one or more liposomes. I.e., the one or more lipids may be formed of, or may comprise, one or more liposomes.

[0142] The composition may in some embodiments comprise incomplete liposomes. In that sense, their interior space may be accessible from the exterior. An incomplete liposome may be visualised as an incomplete lipid “bubble” wherein there are one or more gaps in the (approximately spherical) lipid bilayer surface. Thus, suitably, the one or more lipids may be formed of, or may comprise, one or more incomplete liposomes.

[0143] Suitably, the one or more lipids may be formed of, or may comprise, one or more incomplete liposomes and / or one or more (complete) liposomes.

[0144] Thus, the present composition may comprise the particles comprising hydrolysable silicon associated with one or more liposomes and / or one or more incomplete liposomes, wherein the API is associated with (especially, bound to) the particles comprising hydrolysable silicon. An amino acid (especially, glycine, arginine and / or tyrosine, such as glycine) may also be associated with the particles comprising hydrolysable silicon.

[0145] The optionally present liposomes or incomplete liposomes, may have a mean diameter in a range of from about 50 nm to about 400 nm, especially about 50 nm to about 200 nm, more especially about 60nm to about lOOnm.

[0146] API may be bound non-covalently to particles comprising hydrolysable silicon that are bound to the surface of one or more liposomes and / or one or more incomplete liposomes. Up to about 10 or 20 % of the total API present in the composition may be bound non-covalently to particles comprising hydrolysable silicon that are, in turn, bound to the surface of one or more liposomes and / or one or more incomplete liposomes. API may be bound non-covalently to particles comprising hydrolysable silicon that are, in turn, in the interior of one or more liposomes and / or one or more incomplete liposomes. At least about 50, 60 or 70 % of total API present in the composition may preferably be bound non-covalently to particles comprising hydrolysable silicon that are in the interior of one or more liposomes and / or one or more incomplete liposomes.

[0147] Optionally, API is bound non-covalently to particles comprising hydrolysable silicon that are in the interior of one or more liposomes and / or one or more incomplete liposomes; and API is bound non-covalently to particles comprising hydrolysable silicon that are bound to the surface of one or more liposomes and / or one or more incomplete liposomes.

[0148] Optionally, the composition may be free or substantially free of liposomes; and / or may be free or substantially free of incomplete liposomes.

[0149] Optionally, the one or more lipids may be formed of, or may comprise, one or more lipid monolayers. Optionally, the one or more lipids may be or comprise one or more micelles or incomplete micelles. It will be understood that micelles have similar characteristics to liposomes, except that micelles’ walls are formed of lipid monolayer; whereas liposomes’ walls are formed of lipid bilayer. Thus, a micelle may refer to a vesicle having at least one lipid monolayer, which may be approximately spherical in shape. A micelle, similarly to a liposome, may be visualised as a lipid “bubble” surrounding an interior space. The interior space may be a hydrophilic environment.

[0150] Thus, the present composition may comprise the particles comprising hydrolysable silicon associated with one or more micelles and / or one or more incomplete micelles, wherein the API is associated with (especially, bound to) the particles comprising hydrolysable silicon.

[0151] The optionally present micelles may have a mean diameter in a range of from about 50 nm to about 400 nm, especially about 50 nm to about 200 nm, more especially about 60nm to about lOOnm. API may be bound non-covalently to particles comprising hydrolysable silicon that are bound to the surface of one or more micelles and / or one or more incomplete micelles. Up to about 10 or 20 % of the total API present in the composition may be bound non-covalently to particles comprising hydrolysable silicon that are, in turn, bound to the surface of one or more micelles and / or one or more incomplete micelles.

[0152] API may be bound non-covalently to particles comprising hydrolysable silicon that are, in turn, in the interior of one or more micelles and / or one or more incomplete micelles. At least about 50, 60 or 70 % of the total API present in the composition may preferably be bound non-covalently to particles comprising hydrolysable silicon that are in the interior of one or more micelles and / or one or more incomplete micelles.

[0153] Optionally, API is bound non-covalently to particles comprising hydrolysable silicon that are in the interior of one or more micelles and / or one or more incomplete micelles; and API is bound non-covalently to particles comprising hydrolysable silicon that are bound to the surface of one or more micelles and / or one or more incomplete micelles.

[0154] That said, the composition may be free or substantially free of micelles; and / or may be free or substantially free of incomplete micelles.

[0155] The one or more lipids may be formed of, or may comprise, one or more lipid globules, each globule optionally being surrounded by a layer of surfactants. Lipid globules do not enclose an interior space or cavity. Instead, they are solidly formed, or substantially solidly formed, of lipid, in which other components, such as the particles comprising hydrolysable silicon to which are bound API molecules, may be dispersed. Thus, the globules’ interior may be studded with the particles comprising hydrolysable silicon; with API molecules, in turn, being bound (non-covalently) to the particles comprising hydrolysable silicon. Additionally or alternatively (preferably, additionally), the particles comprising hydrolysable silicon, to which are (non- covalently) bound API molecules, may be bound to the surface of one or more lipid globules. Thus, preferably, the present composition comprises the particles comprising hydrolysable silicon associated with (especially, dispersed within and / or bound onto the surface of) one or more (solid or substantially solid; i.e., non-hollow) lipid globules, wherein the API is associated with (especially, bound to) the particles comprising hydrolysable silicon.

[0156] The optionally present lipid globules may have a mean diameter in a range of from about 50 nm to about 400 nm, especially about 50 nm to about 200 nm, more especially about 60nm to about lOOnm.

[0157] API may be bound non-covalently to particles comprising hydrolysable silicon that are bound to the surface of one or more lipid globules. Up to about 10 or 20 % of total API present in the composition may be bound non-covalently to particles comprising hydrolysable silicon that are, in turn, bound to the surface of one or more lipid globules.

[0158] API may be bound non-covalently to particles comprising hydrolysable silicon that are, in turn, in the interior of one or more lipid globules. At least about 50, 60 or 70 % of total API present in the composition may preferably be bound non-covalently to particles comprising hydrolysable silicon that are in the interior of one or more lipid globules.

[0159] Preferably, API is bound non-covalently to particles comprising hydrolysable silicon that are in the interior of one or more lipid globules; and API is bound non- covalently to particles comprising hydrolysable silicon that are bound to the surface of one or more lipid globules.

[0160] The particles comprising hydrolysable silicon may coalesce into aggregates of particles comprising hydrolysable silicon, for example as shown in the transmission electron microscope (TEM) images of Figure 17. Thus, the described composition may comprise aggregates (especially, chains) of the particles comprising hydrolysable silicon.

[0161] As used herein, the term “aggregate of particles comprising hydrolysable silicon” may refer to a cluster of particles wherein nearest-neighbour particles are in contact with each other. Such clusters may have varying configurations, such as substantially spherical clusters of particles and / or chains of particles. Particularly preferred are configurations comprising or consisting of chains of particles.

[0162] Thus, the composition may comprise one or more aggregates of particles comprising hydrolysable silicon. Preferably, the aggregates comprise one or more chains of the particles.

[0163] There may, for example, be present at least about 2, 3 or 4 chains on average per aggregate.

[0164] The one or more aggregates may comprise one or more branched chains of the particles. Thus, the one or more aggregates may be formed of, or may comprise, branches formed from chains of the particles. There may, for example, be present at least about 2, 3 or 4 branches per aggregate.

[0165] The one or more aggregates of particles comprising hydrolysable silicon may be associated with the one or more lipids, for example embedded in lipid and / or attached to the surface of lipid.

[0166] The one or more aggregates of particles comprising hydrolysable silicon may be associated with one or more lipid structures described herein, for example embedded within and / or attached to one or more lipid structures.

[0167] As described herein, the one or more lipid structures may be formed of or may comprise one or more of: micelles, incomplete micelles, liposomes, incomplete liposomes and lipid globules. Thus, the one or more aggregates of particles comprising hydrolysable silicon may be associated with (for example, embedded in and / or attached to the surface of) one or more of lipid micelles, incomplete lipid micelles, liposomes, incomplete liposomes and lipid globules.

[0168] In particular, the one or more aggregates of particles comprising hydrolysable silicon may be embedded in or attached to the surface of one or more of: liposomes; incomplete liposomes; and lipid globules.

[0169] The ratio of the longest dimension of an aggregate to the longest dimension of a lipid structure may on average be about 1 :5 to 5 :1, especially about 1 :3 to 3 :1; especially when the lipid structure is or comprises liposomes and / or lipid globules and the one or more aggregates are embedded therein or attached to the surface thereof. This may be measured, for example, by TEM as shown in Figure 17.

[0170] The average longest dimension of an aggregate may be about 50 nm to about 500 nm, especially about 50 nm to about 200 nm, such as about 50 to about 150 nm, such as when measured by TEM. In particular, the one or more aggregates may be or comprise one or more chains of particles, with the average length of a chain being about 50 nm to about 500 nm, especially about 50 nm to about 200 nm, such as about 50 to about 150 nm. The average cross-sectional diameter of a chain may be about 5 nm to about 50 nm.

[0171] The ratio of the longest dimension of an individual particle comprising hydrolysable silicon to the longest dimension of a lipid structure may on average be in a range of from about 1 : 100 to about 1 :2, especially about 1 : 100 to about 1 :5, more especially about 1 : 100 to about 1 :9; especially when the lipid structure is or comprises liposomes and / or lipid globules and the one or more aggregates are embedded therein or attached to the surface thereof. This may be measured, for example, by TEM as shown in Figure 17.

[0172] When the one or more aggregates are present, the average (e.g., mean) diameter of a particle may preferably be about 1 nm to about 50 nm. Additionally or alternatively, the particles may be porous and may have an average (e.g., mean) pore diameter of about 0.1 to about 5 nm, such as about 2 nm. In turn, API may be bound (non-covalently) to one or more of the particles in the aggregates.

[0173] Thus, the one or more aggregates may bind API. When the one or more aggregates are or comprise chains of the particles, such chains may extend into the interior of the lipid structure(s) (especially, into liposomes, incomplete liposomes, and / or lipid globules). In this way, the chains may provide a route for the API (especially, when the API is miRNA or mRNA, most especially miRNA) to be better encapsulated into the lipid. Without wishing to be bound by theory, it is thought that this may shield the API (especially, when the API is miRNA or mRNA, most especially miRNA) from degradation, especially by shielding it from enzymes (especially, in vivo) and preventing or reducing API molecules being available to react with water molecules. It is thought that as the silicon particles degrade over time, API may be released, thus enabling protection of the API until it reaches a target site for release.

[0174] The presence of API may itself facilitate aggregation of the particles. For example, nucleic acid APIs (such as miRNA or mRNA, especially miRNA) have a negative charge (due to their phosphate backbone), which may induce aggregation of the particles. Additionally or alternatively, the presence of Si-0 species on the particles’ surface may induce particle-particle interactions to facilitate aggregation.

[0175] Thus, preferably, the one or more aggregates are or comprise one or more chains of the particles comprising hydrolysable silicon, wherein API (especially, mRNA or miRNA, most especially miRNA) is bound to the particles, and the one or more chains extend into the interior of the lipid structure(s), especially, into liposomes, incomplete liposomes, or lipid globules. This may provide a stabler environment for the API (especially, mRNA or miRNA, most especially miRNA). Additionally or alternatively, it may enable increased API (especially, mRNA or miRNA, most especially miRNA) uptake by the lipid structures, compared to no such aggregate(s) being present. This is in contrast to conventional liposomal delivery vehicles, for which inefficient API uptake may be a problem; for example, it is thought that up to about 80 % of conventional liposomal delivery vehicles formulated into commercially available therapeutic compositions may be “empty” of API.

[0176] Optionally, substantially no particles are present as isolated particles; instead, all particles are present in aggregates.

[0177] A structure formed by the composition (e.g., a structure wherein one or more aggregates of the particles comprising hydrolysable silicon are embedded in or attached to the surface of one or more of: liposomes; incomplete liposomes; and substantially solid lipid globules) optionally exhibits surface roughness. Roughness may be measured in accordance with ISO standard ISO 4287:1997. It may be quantified, for example, by reference to Ra, a one-dimensional average (with units of nm) of profile height deviations from a mean line drawn across a surface.

[0178] Without wishing to be bound by theory, it is thought that such surface roughness (and / or surface charge, quantifiable as zeta potential) may assist with avoiding the detrimental accumulation of the API in the liver; and / or promoting accumulation of the API in the pancreas and / or spleen.

[0179] Amino acid s)

[0180] The composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) an active pharmaceutical ingredient (API), may further comprise one or more amino acids.

[0181] In preferred embodiments, the amino acid(s) may comprise or consist essentially of one or more of glycine, tyrosine and arginine, especially glycine, or a combination of glycine with arginine, or a combination of glycine with tyrosine.

[0182] The term “amino acid” may suitably be used to refer to any artificial or naturally occurring organic compound containing an amine (-NH2) and carboxyl (-COOH) functional group. It includes a, P, y and 6 amino acids. It includes an amino acid in any chiral configuration. The amino acid may, especially, be a naturally occurring a amino acid. It may be a proteinogenic amino acid or a non-proteinogenic amino acid (such as carnitine, levothyroxine, hydroxyproline, ornithine or citrulline).

[0183] The one or more amino acids may help stabilise the silicon particles themselves. In vivo, the one or more amino acids may help to modulate the rate of hydrolysis of the silicon, such that the silicon hydrolyses to bioavailable orthosilicic acid (OSA) degradation product; rather than insoluble polymeric hydrolysis products. In this way, the one or more amino acids may complement the function of the one or more lipids of the present disclosure. Controlling the rate of hydrolysis of the silicon may influence the rate of release of API associated with the silicon. Controlling the rate of API release may modulate the length of the time period during which protection of the API is sustained, especially concerning protection in vivo in the presence of various bodily fluids. Thus, more API may be delivered to a target cell in a given time period, than for an otherwise identical composition.

[0184] Without wishing to be bound by theory, while some amino acids are investigated in the Examples hereinbelow, the mechanism by which the one or more amino acids act may be due to properties of the class of amino acids, such as their behaviour in response to charge-charge interactions; and may thus be generalisable beyond the exemplified amino acids. Thus, different amino acids may be used in different amounts to achieve a given zeta potential and / or a given rate of API release and / or a given rate of hydrolysis of the silicon.

[0185] Additionally or alternatively, amino acids which are neutral or positively charged at physiological pH (about pH 7.4), such as tyrosine or arginine, may stabilise negatively charged APIs (e.g. nucleic acids such as mRNA). Meanwhile, amino acids which are neutral or negatively charged at physiological pH (about pH 7.4) may stabilise positively charged APIs. Nonetheless, the interplay of charge-based and / or other interactions (for example, steric interactions) resulting from the combination of Si, lipid(s) and amino acid(s) may be such that amino acid(s) which are positively charged at physiological pH may help to stabilise positively charged APIs, or amino acid(s) which are negatively charged at physiological pH may help to stabilise negatively charged APIs. The ratio by weight of total lipids (e.g., where the lipids consist of DOTAP and DOPE, the combined weight of DOTAP and DOPE) to total amino acids (e.g., where the amino acids consist of arginine and glycine, the combined weight of arginine and glycine) may be in a range of from about 40: 1 to about 5: 1; such, for example, as about 32: 1.

[0186] Non-reducing disaccharide

[0187] Additionally or alternatively, the composition may comprise one or more nonreducing disaccharides, especially trehalose, as exemplified, for instance, by certain of the compositions of the Examples hereinbelow. The ratio by weight of the one or more lipids (i.e. total lipid components) to non-reducing disaccharide may be in a range of from about 20: 1 to about 1 :1; such, for example, as about 16: 1.

[0188] Cannabinoids (especially, CBD and delta-9 THC)

[0189] Additionally or alternatively, the composition may comprise one or more cannabinoids, such as one or more of THC-acid (THCA); CBD-acid (CBDA); cannabigerol (CBG); cannabigerolic acid (CBGA); cannabichromene (CBC); cannabichromenolic acid (CBCA); cannabichromevarin (CBCV); cannabichromevarinolic acid (CBCV A); cannabidivarin (CBDV); cannabidivarinolic acid (CBDVA); cannabinol (CBN); delta-9 tetrahydrocannabinol (delta-9 THC) and cannabidiol (CBD); especially, delta-9 tetrahydrocannabinol and / or cannabidiol, as exemplified, for instance, by certain of the compositions of the Examples hereinbelow. The ratio by weight of the one or more lipids (i.e. total lipid components) to one or more cannabinoids (i.e., total cannabinoid components) may be in a range of from about 200: 1 to about 10: 1; such, for example, as about 160: 1. The ratio by weight of silicon to the one or more cannabinoids (i.e., total cannabinoid components) may be in a range of from about 20: 1 to 1 : 1, such, for example, as about 10: 1. Examples of compositions having such ratios include 2C-LBSi-CBD-RE0.1 and 2C-LBSi-THC-RE0.1 of the Examples hereinbelow. Other advantageous components

[0190] Optionally, the composition further comprises one or more other advantageous components.

[0191] One or more transfection reagents may be present, especially one or more cationic polymers, such as polyethyleneimine, especially branched polyethyleneimine (bPEI) or a derivative thereof. The one or more transfection reagents may assist with endocytosis for the delivery of an API into a cell.

[0192] Optionally when the one or more lipids comprise a cationic lipid (e.g. DOTAP), the ratio by weight of cationic lipid to transfection reagent is in a range of from about 150: 1 to about 50: 1, especially about 100: 1 to about 70: 1. Examples of compositions having such a ratio include 2C-LBSi-BPEI10-Tyr-RE0.1 with 0.5% trehalose, of the Examples below.

[0193] Optionally when the one or more lipids comprise a phospholipid (e.g. DOPE), the ratio by weight of cationic lipid to transfection reagent is in a range of from about 150: 1 to about 50: 1, especially about 100: 1 to about 70: 1. Examples of compositions having such a ratio include 2C-LBSi-BPEI10-Tyr-RE0.1 with 0.5% trehalose, of the Examples below.

[0194] One or more PEGylated non-lipid molecules may be present, such as one or more PEGylated amines, especially DTA-PEG2000. These may optionally be in addition to, or may replace, the one or more PEGylated lipids described herein and may have a similar function.

[0195] Optionally when the one or more lipids comprise cationic lipid (e.g. DOTAP), the ratio by weight of cationic lipid : PEGylated non-lipid molecule(s) may be in a range of from about 50: 1 to about 2: 1; such, for example, as about 10: 1 to about 2: 1. Examples of compositions having such a ratio include 3ISC30.70-P159.3-AESi-SA- RE0.1 and 3ISC30.70-P159.3-LBSi-SA-RE0.1 of the Examples below. When the one or more lipids comprise phospholipid (e.g., DOPE), the ratio by weight of phospholipid: PEGylated non-lipid molecule(s) may be in a range of from about 50: 1 to about 2: 1; such, for example, as about 10: 1 to about 2: 1. Examples of compositions having such a ratio include 3ISC30.70-P159.3-AESi-SA-RE0.1 and 3ISC30.70-P159.3-LBSi-SA-RE0.1 of the Examples below.

[0196] Ortho silicic acid (OSA) or a derivative thereof may preferably be present, such, for example, as in the form of choline-stabilised OSA (“Ch-OSA”). This may assist with lipid-Si binding.

[0197] Optionally when the one or more lipids comprise cationic lipid (e.g. DOTAP), the ratio by weight of cationic lipid : ortho silicic acid (or a derivative thereof, especially Ch-OSA) may be in a range of from about 10:1 to about 1: 1; such, for example, as about 5: 1 to about 2: 1. Examples of compositions having such a ratio include 2C-LBSi-Ch- OSA0.3-Arg-Gly-SA-RE0.1 and 2C-LBSi-Ch-OSA0.3-Tyr-Gly-RE0.1 of the Examples below.

[0198] When the one or more lipids comprise phospholipid (e.g., DOPE), the ratio by weight of phospholipid: ortho silicic acid (or a derivative thereof, especially Ch-OSA) may be in a range of from about 10:1 to about 1 : 1; such, for example, as about 5:1 to about 2: 1. Examples of compositions having such a ratio include 2C-LBSi-Ch- OSA0.3-Arg-Gly-SA-RE0.1 and 2C-LBSi-Ch-OSA0.3-Tyr-Gly-RE0.1 of the Examples below.

[0199] One or more sialic acids may optionally be present, for example to supplement targeting of the spleen and / or of the pancreas. Additionally or alternatively, sialic acid may assist in prolonging circulation of the composition in vivo.

[0200] Optionally when the one or more lipids comprise cationic lipid (e.g. DOTAP), the ratio by weight of cationic lipid: sialic acid may be in a range of from about 50:1 to about 1 : 1; such, for example, as about 40:1 to about 20: 1. Examples of compositions having such a ratio include 3C-SiAEN-RE0.1, 3C-SiLB-RE0.1, 3ISC30.70-P159.3- AESi-SA-RE0.1, 3ISC30.70-P159.3-LBSi-SA-RE0.1 and 2C-LBSi-Ch-OSA0.3-Arg- Gly-SA-REO. lof the Examples below.

[0201] When the one or more lipids comprise phospholipid (e.g., DOPE), the ratio by weight of phospholipid: sialic acid may be in a range of from about 50: 1 to about 1 : 1; such, for example, as about 40: 1 to about 20: 1. Examples of compositions having such a ratio include 3C-SiAEN-RE0.1, 3C-SiLB-RE0.1, 3ISC30.70-P159.3-AESi-SA- RE0.1, 3ISC30.70-P159.3-LBSi-SA-RE0.1 and 2C-LBSi-Ch-OSA0.3-Arg-Gly-SA- RE0. lof the Examples below.

[0202] The one or more other advantageous components may be or comprise nicotinamide adenine dinucleotide (NAD) or a derivative thereof. Optionally, the ratio by weight of the one or more lipids (i.e., total lipid components) to the NAD may be in a range of from about 130: 1 to about 30: 1, such as about 80: 1.

[0203] Additionally or alternatively, the composition may comprise a flavanol, such as quercetin or a derivative thereof. Optionally, the ratio by weight of the one or more lipids (i.e., total lipid components) to the quercetin may be in a range of from about 130: 1 to about 30: 1, such as about 80: 1.

[0204] APIs

[0205] It will be understood that the composition of the present disclosure may be described as a pharmaceutical composition, being for the delivery of an active pharmaceutical ingredient (API).

[0206] Most preferably, the API is or comprises (especially, is) miRNA.

[0207] The API may, for example, be a fragile API. As used herein, the terms “fragile APIs” and “reactive APIs” may be interchangeable and may refer to APIs which (i) have a half-life upon storage in aqueous solution at about 25 °C of at most one week, measurable by NMR or by GC-MS; and / or (ii) have a half-life in vivo of under about an hour, measurable by assay of a biological sample. The API may be any pharmaceutically active compound; thus, for example, it will be understood that the term “API” encompasses pro-drugs. The API may be a protein. The API may especially be a nucleic acid, more especially miRNA, siRNA or mRNA. As described, it is especially preferred that the API is or comprises miRNA.

[0208] The API may be for administration by injection, orally, intranasally or topically; especially, by injection or orally.

[0209] The ratio by weight of total lipid (e.g., where the lipids consist of DOPE and DOTAP, the weight of total lipid is the weight of the DOPE and the weight of the DOTAP, combined) to total API may be about 5: 1 to about 100: 1, especially about 5:1 to about 50: 1. Examples include a ratio by weight of total lipid to API of about 12: 1, about 24: 1 and about 48: 1.

[0210] APIs for the pancreas

[0211] The API may be or comprise an API for the pancreas. The API may be or comprise an API for the prevention or treatment of a pancreatic disease or disorder. The Examples below show targeting of the pancreas by compositions in accordance with the present disclosure, so that the targeting of the pancreas is especially contemplated herein.

[0212] The API may be or comprise an API for the prevention or treatment of acute or chronic pancreatitis.

[0213] The API may be or comprise one or more steroids, one or more analgesics and / or one or more antibiotics.

[0214] The API may be or comprise one or more proteins, especially one or more enzymes, such as enzymes for supporting pancreatic function.

[0215] The API may be or comprise an API for the prevention or treatment of pancreatic cancer, as is preferred and described herein. The API may be or comprise an API for the prevention or treatment of pancreatic cysts.

[0216] The API may be or comprise an API for the prevention or treatment of an ampullary or periampullary disease or disorder.

[0217] The API may be or comprise an API configured to target the pancreas. The API may be or comprise an API configured to target one or more tissues of the pancreas. The API may be or comprise an API configured to target one or more cells of the pancreas. While the API may target one or more cells of the pancreas, a therapy provided by the API may nevertheless have a systemic effect, for example the modulation of insulin secretion by beta cells, by the API, may prevent, mitigate or treat type 2 diabetes.

[0218] The API may be or comprise an API configured to target the beta cells of the pancreas. The API may be or comprise an API configured to modulate (such as to promote) the secretion of insulin by beta cells.

[0219] The API may be or comprise one or more glucagon-like peptide- 1 (GLP-1) agonists. The API may be or comprise one or more of dulaglutide, exenatide, liraglutide, lixisenatide, semaglutide and derivatives thereof, especially semaglutide and derivatives thereof.

[0220] The API may be for the prevention or treatment of diabetes, such as type 1 or type 2 (especially, type 2) diabetes. Thus, the API may be or comprise one or more antidiabetic APIs, such as one or more glucagon-like peptide- 1 (GLP-1) derivatives, especially semaglutide or derivatives thereof.

[0221] The API may be or comprise one or more immunomodulators, especially one or more antibodies targeting CD-3, such as the CD-3 monoclonal antibody teplizumab. The API may be or comprise one or more neurotransmitters for the treatment of beta cells, such as gamma aminobutyric acid (GABA), dopamine and derivatives thereof.

[0222] The API may be or comprise one or more endoplasmic reticulum chaperones, such as tauroursodeoxycholic acid (TUDCA), deoxycholic acid, ursodeoxycholic acid (UDCA), glycoursodeoxy cholic acid or derivatives (e.g., salts) thereof.

[0223] The API may be or comprise one or more anti-obesity APIs, such as one or more GLP-1 derivatives, especially semaglutide or derivatives thereof.

[0224] Nucleic acid APIs

[0225] The API may preferably be or comprise a nucleic acid, especially RNA (linear or circular RNA). The RNA may be small interfering RNA (siRNA), small activating RNA (saRNA), small hairpin RNA (shRNA), transfer RNA (tRNA), microRNA (miRNA) or messenger RNA (mRNA), especially mRNA (e.g., mRNA that encodes a protein of a pathogenic organism), siRNA or miRNA, most especially miRNA.

[0226] Other nucleic acids for use in accordance with the present disclosure include: double- and single-stranded DNA; DNA:RNA hybrids; peptide:DNA hybrids; and peptide:RNA hybrids.

[0227] A method disclosed herein may comprise a step (in vivo or in vitro, preferably in vivo) of transfecting a human cell with the nucleic acid API, using the composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) the nucleic acid API.

[0228] RNA and DNA may be naturally occurring or chemically modified to enhance their therapeutic properties, such as enhanced activity, increased serum stability, reduced off-targeting and lower immunological activation. Chemical modifications to RNA and DNA may include any modifications commonly known in the art. As used herein, the term “naturally occurring” means of natural human or animal origin. It will be understood that a molecular structure that is the same as a naturally occurring molecular structure may nevertheless be synthesized in vitro, such as when mRNA is synthesized by in vitro transcription (IVT).

[0229] Thus, as used herein, the terms nucleic acid, DNA and RNA also include known types of modifications, for example, labels which are known in the art, methylation, “caps”, substitution of one or more of the naturally occurring nucleotides with an analogue, internucleotide modifications such as, for example, those with uncharged linkages (e.g., methyl phosphonates, phosphotriesters, phosphoramidates, carbamates, etc.), with negatively charged linkages (e.g., phosphorothioates, phosphorodithioates, etc.), and with positively charged linkages (e.g., aminoalklyphosphoramidates, aminoalkylphosphotriesters), those containing pendant moieties, such as, for example, proteins (including nucleases, toxins, antibodies, signal peptides, poly-L-lysine, etc.), those with intercalators (e.g., acridine, psoralen, etc.), those containing chelators (e.g., metals, radioactive metals, boron, oxidative metals, etc.), those containing alkylators, those with modified linkages (e.g., alpha anomeric nucleic acids, etc.), as well as unmodified forms of the polynucleotide or oligonucleotide.

[0230] Similarly, as used herein, the terms “nucleoside” and “nucleotide” will include those moieties which contain not only the known purine and pyrimidine bases, but also other heterocyclic bases which have been modified. Such modifications include methylated purines or pyrimidines, acylated purines or pyrimidines, or other heterocycles. Modified nucleosides or nucleotides will also include modifications on the sugar moiety, e.g., wherein one or more of the hydroxyl groups are replaced with a halogen, an aliphatic group, or are functionalized as ethers, amines, or the like. Other modifications to nucleotides or polynucleotides involve rearranging, appending, substituting for, or otherwise altering functional groups on the purine or pyrimidine base which form hydrogen bonds to a respective complementary pyrimidine or purine, e.g., isoguanine, isocysteine, and the like. In some embodiments, the oligonucleotides and / or probes include at least one, two, three or four modified nucleotides.

[0231] In some embodiments, the nucleic acids such as the RNAs disclosed herein include one or more universal bases. As used herein, the term “universal base” refers to a nucleotide analogue that can hybridize to more than one nucleotide selected from A, U / T, C, and G. In some embodiments, the universal base can be selected from the group consisting of deoxyinosine, 3-ntiropyrrole, 4-nitroindole, 6-nitroindole, 5-nitroindole.

[0232] The term “siRNA” may suitably be used to refer to small interfering RNA, comprising RNA molecules which operate within the RNA interference (RNAi) pathway. siRNA is sometimes known as short interfering RNA or silencing RNA. The siRNA may be double stranded. The siRNA may have a length in a range of from about 5 to about 50 base pairs, especially about 10 to about 40 base pairs, more especially about 15 to about 30 base pairs.

[0233] The term “mRNA” may suitably be used to refer to messenger RNA for the synthesis of protein(s). It may encompass mRNA comprising a 5-prime cap and / or a poly-adenylated terminus. Alternatively, one or both of those features may be absent. Typically, the mRNA may be single stranded. The coding region of the mRNA may be at least about 100, especially at least about 500, more especially at least about 1000 bases in length.

[0234] The mRNA may encode an antigen, thereby providing a composition which is a vaccine. The antigen may be a bacterial, parasitic or fungal antigen. The antigen may be a viral antigen, especially a viral antigen of one of the viral diseases described hereinbelow; more especially an antigen of a respiratory virus, for example an antigen of SARS-CoV-2, for example an antigen deriving from the spike protein of SARS- CoV-2.

[0235] The mRNA may encode an allergen (including but not limited to one or more nut allergens; which in turn include, but are not limited to: one or more seed storage proteins, such as vicilins, legumins, albumins; one or more plant defence related proteins; and one or more profilins).

[0236] The mRNA may encode a protein that modulates an immune, autoimmune, or inflammatory disease (including, but not limited to, lupus, atherosclerosis, chronic obstructive pulmonary disease, inflammatory bowel disease, multiple sclerosis, psoriasis, a rheumatic disease, uveitis, atopic dermatitis, and pulmonary fibrosis). The mRNA may encode a tumour-specific antigen. As used herein, the term tumour-specific antigen may refer to an antigen that arises, in one or more malignant cancer cells, from non-synonymous somatic mutation (leading to a neoantigen) or viral- integrated mutation (leading to an onco-viral antigen). Tumour-specific antigens may thus refer to antigens that are completely absent from (not expressed by) non-cancerous (healthy, normal) cells.

[0237] The mRNA may encode a tumour-associated antigen. As used herein, the term tumour-associated antigen may refer to an antigen that is over-expressed in a malignant cancer cell, compared to a non-cancerous (healthy, normal) cell, for example due to genetic amplification or post-translational modifications. The term tumour-associated antigen may encompass overexpressed antigens (which term may refer to proteins that are moderately expressed in non-cancerous (healthy, normal) cells, but expressed abundantly in malignant cancer cells); differentiation antigens (which term may refer to proteins that are selectively expressed by the cell lineage from which the malignant cells evolved, an example being prostate-specific antigen); and cancer-germline antigens (which term may refer to antigens that are normally limited to reproductive tissues, but which are aberrantly expressed in a malignant cancer cell; for example, melanoma antigen family A3 (MAGE-A3); New York Esophageal Squamous Cell Carcinoma- 1 Antigen (NY-ESO-1); and Preferentially Expressed Antigen in Melanoma (PRAME)).

[0238] The mRNA may encode multiple proteins, thereby providing more effective pharmacological activity. The mRNA may encode multiple antigens, especially multiple viral antigens.

[0239] The mRNA may additionally encode an adjuvanting protein. An adjuvant may additionally or alternatively be provided as a further component of the composition in addition to the API.

[0240] The term “miRNA” may suitably be used to refer to micro RNA; especially, micro RNA comprising single-stranded RNA molecules configured to silence mRNA. Thus, miRNA is typically configured to regulate gene expression post-transcriptionally; i.e., miRNA typically operates in pathways for the post-transcriptional regulation of gene expression. miRNA may be configured to base-pair to complementary sequences in mRNA molecules. This may enable the miRNA to silence said mRNA molecules, such as by cleavage of an mRNA strand or shortening of an mRNA poly-A tail (shortening of the tail leading to mRNA destabilisation and degradation). As described, miRNA may typically be single stranded. miRNA may have a length in a range of from about 5 to about 50 base pairs, especially about 10 to about 40 base pairs, more especially about 15 to about 30 base pairs. miRNA expression may be dysregulated in cancer. In particular, aberrant downregulation of miRNA in cancer may result in pathological overexpression of oncogenes. Accordingly, miRNA replacement is a promising therapeutic strategy. miRNA may therefore be configured as a tumour suppressor. As an API, therefore, miRNA may mitigate, treat or prevent cancer.

[0241] Additionally or alternatively, miRNA may sensitise cancer cells to one or more other therapies, especially one or more chemotherapeutic agents. miRNA may mitigate, treat or prevent the resistance of cancer cells to one or more other therapies, especially one or more chemotherapeutic agents.

[0242] It will be understood that miRNA is not siRNA.

[0243] For example, a given siRNA is configured to inhibit the expression of one specific mRNA. In sharp contrast, a given miRNA is configured to regulate the expression of multiple mRNAs. This may be because miRNA typically acts by binding to 3' untranslated regions (UTRs) of target mRNA with imperfect complementarity, causing translational repression or mRNA degradation. The requirement for a merely imperfect match may mean that a single miRNA can regulate multiple different genes. Whereas, siRNA typically binds to target mRNA with near perfect complementarity, leading to RNA-induced silencing complex (RlSC)-mediated cleavage and degradation of the target mRNA. Thus, siRNA may be more selective in its activity, than miRNA. Typically in vivo, when miRNA associates, as a duplex, with an RNA-induced silencing complex (RISC), forming a miRISC complex, the miRNA duplex is unwound, releasing and discarding a passenger strand (sense strand). This is unlike typical processing of siRNA, in which the argonaute 2 protein (AGO2) of a RISC causes the cleavage of the passenger strand of siRNA.

[0244] Furthermore, miRNA synthesis in vivo typically involves transcription by RNA polymerase II to form primary miRNA (pri-miRNA), which is then processed in the nucleus into a precursor miRNA (pre-miRNA) by the Drosha-DGCR8 complex, and further processed in the cytoplasm by Dicer into a mature miRNA duplex. In contrast, siRNA synthesis in vivo typically involves Dicer processing of double-stranded RNA (dsRNA) directly into siRNA duplexes, which are then unwound to form singlestranded siRNAs that are loaded into the RISC.

[0245] It will also be understood that miRNA is not mRNA. mRNA serves as a template for protein synthesis, in translation. miRNA does not encode protein; rather, miRNA regulates mRNA expression. Another difference lies in the implementation of miRNA, compared to mRNA, in cancer therapy. mRNA cancer therapy seeks to stimulate immune cells thereby to generate an anti-tumour immune response (wherein the immune cells proceed to attack the cancer cells). In contrast, miRNA acts directly on cancer cells (without the intermediary of immune cells).

[0246] Chemotherapeutic agent

[0247] As used herein, the term chemotherapeutic agent refers to an active pharmaceutical agent (API; i.e., a drug) for the treatment or mitigation of cancer. Administration of one or more chemotherapeutic agents to a patient in need thereof is known in the art as cancer chemotherapy, or simply as chemotherapy.

[0248] A chemotherapeutic agent may typically be or comprise a cytotoxic API to destroy or inhibit the growth and division of malignant cancerous cells.

[0249] The term neurotoxic chemotherapeutic agent refers to a chemotherapeutic agent that exhibits toxicity towards cells of the nervous system. Examples of neurotoxic chemotherapeutic agents include, without limitation: taxanes such as paclitaxel (taxol) and docataxel; platinum-containing chemotherapeutic agents such as cisplatin, carboplatin and oxaliplatin; vinca alkaloids such as vincristine; cytabarine; ifosfamide; bortezomib; and methotrexate. In some embodiments, the cancer chemotherapy received by the patient comprises treatment by one or more platinum-containing chemotherapeutic agents. In some embodiments, the one or more chemotherapeutic agents are selected from one or more of: cisplatin, paclitaxel, docetaxel, vincristine, oxaliplatin, and bortezomib. Neurotoxic chemotherapeutic agents may have especially significant side effects, meaning a patient and their physician decide to discontinue chemotherapy using such agents after a short period of time, such as before completion of a scheduled chemotherapeutic treatment.

[0250] Administering miRNA concomitantly with one or more chemotherapeutic agents (such as one or more neurotoxic chemotherapeutic agents) or in intervals between administration of one or more chemotherapeutic agents (it will be understood that in such intervals, the patient is not administered the one or more chemotherapeutic agents) may sensitise cancer cells to the one or more chemotherapeutic agents and thus enable lower doses of the one or more chemotherapeutic agents to be administered; and / or a longer course of administration of (i.e., treatment with) the one or more chemotherapeutic agents.

[0251] One or more chemotherapeutic agents may be administered to a patient to treat one or more of: ovarian cancer; breast cancer; squamous cell cancer; lung cancer (including small-cell lung cancer and non-small cell lung cancer); adenocarcinoma of the lung; squamous carcinoma of the lung; cancer of the peritoneum; hepatocellular cancer; gastric or stomach cancer (including gastrointestinal cancer); pancreatic cancer; glioblastoma; cervical cancer; liver cancer; bladder cancer; hepatoma; colon cancer; colorectal cancer; endometrial or uterine carcinoma; salivary gland carcinoma; kidney or renal cancer; liver cancer; prostate cancer; vulval cancer; thyroid cancer; hepatic carcinoma; various types of cancer of the head and neck; B-cell lymphoma (including low grade / follicular non-Hodgkin's lymphoma (NHL); small lymphocytic NHL; intermediate grade / follicular NHL; intermediate grade diffuse NHL; high grade immunoblastic NHL; high grade lymphoblastic NHL; high grade small non-cleaved cell NHL; bulky disease NHL; mantle cell lymphoma; AIDS-related lymphoma; Waldenstrom's Macroglobulinemia; chronic lymphocytic leukemia; acute lymphoblastic leukemia; hairy cell leukemia; chronic myeloblastic leukemia; and posttransplant lymphoproliferative disorder.

[0252] In particular, one or more chemotherapeutic agents may be administered to a patient to treat pancreatic cancer, especially pancreatic ductal adenocarcinoma (PDAC).

[0253] Chemotherapy may involve treatment by a single chemotherapeutic agent. Chemotherapy may involve treatment by more than one chemotherapeutic agent.

[0254] Dosage regimes

[0255] Provided herein is a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs, for use in a method of treating cancer in a human patient in need thereof. The composition may be for use with one or more chemotherapeutic agents in a method of treating cancer in a human patient in need thereof.

[0256] Also provided herein is a method of treating or preventing cancer, comprising administering to a human patient in need thereof a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs. The method may further comprise administering one or more chemotherapeutic agents to the patient.

[0257] Factors governing administering one or more chemotherapeutic agents to the patient may include the type of cancer, the extent of cancer, the type(s) of cancer chemotherapy agent(s), the toxicity of the agent(s), and whether the patient and their physician decide to continue or discontinue chemotherapy. More than one different chemotherapeutic agent may be received simultaneously. More than one different chemotherapeutic agent may be received sequentially.

[0258] A patient may receive a single dose of a chemotherapeutic agent. A patient may receive more than one dose of a chemotherapeutic agent. The composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs may be administered before a patient receives their first dose of one or more chemotherapeutic agents. As described herein, the miRNA may sensitise cancer cells to one or more chemotherapeutic agents, so that a lower dose of chemotherapy may be administered.

[0259] Additionally or alternatively, the composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs may be administered after a patient receives their first dose of one or more chemotherapeutic agents.

[0260] It will be understood that as used herein, the term “first dose” refers to the first ever dose administered to a particular patient in respect of a particular cancer. In some circumstances, the patient may have been administered one or more chemotherapeutic agents before, such as for a different cancer, or for the same cancer, which has now become relapsed or refractory.

[0261] Accordingly, it will be appreciated that the cancer may be relapsed or refractory.

[0262] One or more doses of the chemotherapeutic agent(s) may be received over a period of time.

[0263] For example, one or more doses may be received in one or more cycles of chemotherapy over a period of time.

[0264] As used herein, the term cycle of chemotherapy may refer to: (i) a treatment period during which is administered the one or more chemotherapeutic agents (for example, by IV drip over a period in a range of from about 1 to about 2 days); together with (ii) a rest period for the entire duration of which no chemotherapeutic agents are administered to the patient. Thus, a single treatment period (i) and a single rest period (ii) may together make up a single cycle. Cycles may be repeated, such as on a weekly, bi-weekly, or monthly basis. A patient may receive one, two, three, four, five, six or more cycles of chemotherapy.

[0265] The rest period may allow the patient to recover from one or more side effects of the one or more chemotherapeutic agents. During the rest period, the blood plasma level of the or each chemotherapeutic agent(s) in the patient may drop to below 0.5, 0.1 or 0.001 pg / L.

[0266] Preferably, the composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs is administered to the patient when the blood plasma level of the chemotherapeutic agents in the patient is below 0.5, 0.1 or 0.001 pg / L.

[0267] A patient may be administered the composition and one or more chemotherapeutic agents, whether in regular cycles of treatment or sporadically, at certain times within a period of at least about 1 month, at least about 2 months, at least about 3 months, at least about 6 months, at least about 9 months, at least about 1 year, at least about 2 years, at least about 3 years, at least about 4 years, at least about 5 years, or more. Preferably, the composition is administered separately to the one or more chemotherapeutic agents, and when the blood plasma level of the chemotherapeutic agents in the patient is below 0.5, 0.1 or 0.001 pg / L.

[0268] Preferably, the composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs, is administered to the patient in intervals between chemotherapeutic cycles or in one or more rest periods of one or more chemotherapeutic cycles (as defined herein); thus is administered when the patient is not being administered the one or more chemotherapeutic agents.

[0269] Most preferably, the composition is administered to the patient in one or more rest periods of one or more chemotherapeutic cycles.

[0270] The composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs may sensitise cancer cells to the one or more chemotherapeutic agents. Thus, it may enable a lower dose of the one or more chemotherapeutic agents to be administered.

[0271] Additionally or alternatively, the composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) one or more miRNAs may itself have anti-tumour activity, for example by including tumour suppressive miRNA.

[0272] At the same time, delivery of the miRNA itself may present a challenge. The more miRNA that can be delivered to cancer cells, the more tumour suppression may be achieved, whether directly by the miRNA or indirectly by miRNA sensitising cancer cells to chemotherapy.

[0273] Treatment efficiency may be increased by delivering the miRNA in a targeted manner to tumour cells. By targeting miRNA to cancer cells, less miRNA can be used to achieve the same effect on the tumour cells, compared to non-targeted delivery wherein miRNA is wasted elsewhere in the body.

[0274] The composition comprising the (i) one or more lipids and the (ii) particles comprising hydrolysable silicon is thought to provide a delivery vehicle capable of targeting miRNA to cancer cells, and / or to tissues or organs in which cancer cells are present, especially the pancreas and / or spleen.

[0275] Cancer

[0276] The cancer may be or comprise one or more solid tumours.

[0277] The cancer may be or comprise one or more of: ovarian cancer; breast cancer; squamous cell cancer; lung cancer (including small-cell lung cancer and non-small cell lung cancer); adenocarcinoma of the lung; squamous carcinoma of the lung; cancer of the peritoneum; hepatocellular cancer; gastric or stomach cancer (including gastrointestinal cancer); pancreatic cancer; glioblastoma; cervical cancer; bladder cancer; hepatoma; colon cancer; colorectal cancer; endometrial or uterine carcinoma; salivary gland carcinoma; kidney or renal cancer; prostate cancer; vulval cancer; thyroid cancer; hepatic carcinoma; and various types of cancer of the head and neck. The cancer may be or comprise one or more blood cancers, such, for example, as a leukaemia, a lymphoma, or a myeloma.

[0278] The cancer may be or comprise one or more of: acute myeloid leukaemia (AML); chronic myeloid leukaemia (CML); acute lymphoblastic leukaemia (ALL); chronic lymphocytic leukaemia (CLL); non-Hodgkin lymphoma (NHL); Hodgkin lymphoma (HL); or myeloma. Thus, in some implementations, the cancer may be B cell or T cell acute lymphoblastic leukaemia; or large cell lymphoma. Especially, the cancer may be aggressive, relapsed or refractory non-Hodgkin lymphoma, including: diffuse large B cell lymphoma, primary' mediastinal B-cell lymphoma, high grade B- cell lymphoma, transformed follicular lymphoma, mantle cell lymphoma and peripheral T cell lymphoma.

[0279] The cancer may especially be or comprise pancreatic cancer. The Examples below show targeting of the pancreas by compositions in accordance with the present disclosure, so that the prevention or treatment of pancreatic cancer is especially contemplated herein.

[0280] More especially, the cancer may be or comprise exocrine pancreatic cancer, most especially pancreatic ductal adenocarcinoma (PDAC; sometimes also known in the art as invasive pancreatic adenocarcinoma, or simply as pancreatic adenocarcinoma).

[0281] Thus the cancer most especially is or comprises exocrine pancreatic adenocarcinoma.

[0282] The pancreatic cancer may arise in the pancreatic ductal epithelium.

[0283] The pancreatic cancer may be an adenocarcinoma in the head of the pancreas, which sits within the curvature of the duodenum, and wraps around the superior mesenteric artery and vein. Besides pancreatic ductal adenocarcinoma, the exocrine pancreatic cancer may be or comprise an acinar cell carcinoma of the pancreas, a cystadenocarcinoma, a pancreatoblastoma, an adenosquamous carcinoma, a signet ring cell carcinoma, a hepatoid carcinoma, a colloid carcinoma, an undifferentiated carcinoma (optionally, with osteoclast-like giant cells) and / or a solid pseudopapillary tumour.

[0284] The pancreatic cancer may be or comprise a pancreatic neuroendocrine tumour.

[0285] The pancreatic cancer may be characterized by signs and / or symptoms including pain in the upper abdomen or back (often spreading from around the stomach to the back); jaundice (a yellow tint to the whites of the eyes or skin, with or without pain, and possibly in combination with darkened urine; jaundice may result when a cancer in the head of the pancreas obstructs the common bile duct as it runs through the pancreas); unexplained weight loss (either from loss of appetite, or loss of exocrine function resulting in poor digestion); nausea (such, for example, as when a pancreatic tumour compresses neighbouring organs, disrupting digestive processes and making it difficult for the stomach to empty); and constipation.

[0286] The pancreatic cancer may metastasise, i.e., develop secondary malignant growths at a distance from the primary site of cancer. Thus, the pancreatic cancer may be metastatic pancreatic cancer, especially metastatic PDAC (such as metastatic pancreatic cancer that has spread to the spleen).

[0287] The cancer may be or comprise cancer of the spleen. It may be or comprise a splenic tumour that is a lymphoma or a sarcoma.

[0288] Where the cancer comprises cancer of the spleen, it may be due to metastasis of a cancer that first developed elsewhere in the body, such as in the pancreas.

[0289] Thus, the cancer may comprise both pancreatic cancer and cancer of the spleen, such as wherein the cancer has spread from the pancreas to the spleen. The Examples below show targeting of the pancreas and spleen by compositions in accordance with the present disclosure, so that the prevention or treatment of pancreatic and splenic cancer is especially contemplated herein.

[0290] Any cancer described herein may be relapsed (when cancer returns despite successful treatment) and / or refractory (when cancer is resistant to treatment). It will be appreciated that treatment with miRNA-containing compositions described herein, which may have direct tumour suppressive activity and / or sensitise cancer cells to one or more chemotherapeutic agents, may be especially beneficial when the cancer is relapsed and / or refractory.

[0291] The liver; and the detrimental accumulation of an API (such as miRNA) therein

[0292] The liver may be divided into a number of lobes. Each lobe may be sub-divided into a number of hepatic lobules, in which are present hepatocytes. The liver is involved in carbohydrate, protein, amino acid, and lipid metabolism. The liver is also involved in the breakdown (and excretion) of waste products, toxic substances (e.g., by methylation thereof) and, critically, of APIs. Large quantities of blood may pass through the liver and / or be present in it at any given time. Its normal blood volume, including both that in the hepatic veins and that in the hepatic sinuses, may be about 450 ml (in typical adult humans), or almost 10 % of the body's total blood volume. Taken together, these factors mean that the liver may undesirably sequester API molecules and break them down. This may occur before the API has had an opportunity to circulate more widely in the body, especially before the API has had an opportunity to circulate in sufficient quantities and / or for a sufficient time period to a target organ, tissue or cells outside of the liver, such as the pancreas and / or spleen.

[0293] The present inventors have found that the compositions described herein may reduce or prevent the accumulation in the liver of an API as described herein, especially that they may reduce or prevent the detrimental accumulation of the API in the liver; i.e., that they may enable API to “bypass” or “escape” the liver.

[0294] Thus, compositions and methods described herein may be for reducing or preventing the accumulation (i.e., build-up) of the API (especially, miRNA) in (i.e., sequestering of the API by) the liver. This action may be especially useful in the context of cancer. Thus, treating cancer (especially by miRNA administration to a human patient in need thereof) may comprise reducing or preventing the accumulation of an API (especially miRNA) in the liver. It will be understood that some cancers require API delivery to a small area (such as a small single area of tissue) while others require systemic API delivery (such as to multiple tumour-containing organs); across this range of cancers, there remains benefit in API not being sequestered and metabolised by the liver.

[0295] As used herein, the term “detrimental accumulation” may mean accumulation in the liver of an API for therapy of an organ, tissue and / or cell outside the liver; such, in particular, as an API (especially, miRNA) for therapy of the pancreas (especially, pancreatic cancer).

[0296] The accumulation in the liver may be or involve build-up of the API in the liver and break down (metabolism) of the API by the liver. Thus, detrimental accumulation in the liver may be defined or summarised simply as metabolism of the API by the liver. This is detrimental for the reason that it sequesters API from target sites outside the liver.

[0297] Accumulation in the liver may mean that no more than about 10, 20, 30, 40 or 50 % of total API that enters the bloodstream is ever available (especially, available in active form, i.e., before it is metabolised, especially metabolised by the liver) to any organ, tissue or cell outside the liver.

[0298] Accumulation in the liver may mean that the API is sequestered by the liver and is broken down by the liver to the extent that it is substantially unavailable (especially, substantially unavailable in active, unmetabolized form) to any organ, tissue or cell outside the liver.

[0299] Accumulation in the liver may especially mean that the half-life in vivo of the API is reduced. This may be because of the rate at which the API is broken down by the liver. Thus, accumulation in the liver may be characterised by a reduction in the half life in vivo of the API by a factor of two, three, or four.

[0300] Accumulation in the liver may suitably be measured by well-known techniques, such as by positron emission tomography (PET) of radio-labelled API; or optical imaging (e.g., IVIS imaging, as in Example 2 below) of fluorescently labelled API. These techniques may typically be non-invasive (especially, whole-body PET or wholebody IVIS).

[0301] As such, accumulation in the liver may mean that the majority of (or at least about 30, 40, 50, 60 or 70 % of) total API administered (such as administered orally or by injection) to the patient (or, total API that enters the bloodstream of the patient) enters the liver and is there broken down (thus, it does not leave the liver again in the form of functional API), without reaching other tissues; which may be quantified by PET of radio-labelled API or optical imaging (e.g., IVIS) of fluorescently labelled API.

[0302] It will be understood that where % of total API is referred to herein, it may be taken to mean % by weight, based on the weight of total API administered to the patient (or, which enters the bloodstream, if appropriate).

[0303] Thus, reducing or preventing the accumulation of the API in the liver may be or comprise preventing at least about 30, 40, 50, 60 or 70 % of total API administered (such as administered orally or by injection) to the patient from entering (and, especially, from being broken down in) the liver.

[0304] Where appropriate (e.g., if the API is administered orally), reducing or preventing the accumulation of the API in the liver may be or comprise preventing at least about 30, 40, 50, 60 or 70 % of total API that enters the bloodstream of the patient from entering (and, especially, from being broken down in) the liver.

[0305] Reducing or preventing the accumulation of the API in the liver may preferably be or comprise reducing or preventing the accumulation of the API in the liver while promoting accumulation of the API in one or more target organs, one or more target tissues and / or one or more target cells; especially, in a target organ. Reducing or preventing the accumulation of the API in the liver may mean that less (such as about 2x, 3x, 4x or 5x less) API enters the liver, than enters the target organ. Where no API enters the liver, most API may enter the target organ.

[0306] Especially, reducing or preventing the accumulation of the API in the liver may mean that less (such as about 2x, 3x, 4x or 5x less) API enters the liver, than enters the pancreas and / or spleen. Where no API enters the liver, most API may enter the pancreas and / or spleen.

[0307] The pancreas; and promoting accumulation of an API (such as miRNA) therein

[0308] The anatomy of the pancreas may be divided into a head, neck, body, and tail. The head surrounds the superior mesenteric artery and vein. The body is proximate the stomach. The tail is proximate the spleen. Two main pancreatic ducts and a smaller accessory pancreatic duct run through the body of the pancreas.

[0309] The pancreatic islets or islets of Langerhans are regions of the pancreas that contain endocrine cells. Beta cells (p-cells) are endocrine cells located within the pancreatic islets and are responsible for the production and release of insulin and amylin.

[0310] Although the pancreas has an extensive blood supply (with vessels originating as branches of both the coeliac artery and superior mesenteric artery) and freshly ingested or injected API may in some instances reach the pancreas before it reaches the liver, the API may quickly be metabolised by the liver, leading to a reduced steady state concentration developed over time in the blood and a reduced availability of the API to the pancreas. This may make it difficult to trigger API accumulation in the pancreas.

[0311] Compositions and methods in accordance with the present disclosure may be for promoting the accumulation of an API (especially, miRNA) in the pancreas.

[0312] As used herein, the term accumulation in the pancreas may mean accumulation in the pancreas of an API for therapy of a pancreatic tissue and / or cell; such, in particular, as an miRNA for therapy of pancreatic cancer, especially of PDAC; or an API for the therapy of beta cells, thus for the prevention or treatment of diabetes. The accumulation in the pancreas may be or involve build-up of the API in one or more tissues (or cells) of the pancreas and / or treatment of the one or more tissues (or cells) of the pancreas by the API.

[0313] Accumulation in the pancreas may mean that more than about 10, 20, 30 or 50 % of total API that enters the bloodstream is available (especially, available in active form, i.e., before it is metabolised, especially metabolised by the liver) for the treatment of one or more pancreatic tissues and / or cells.

[0314] Accumulation in the pancreas may mean that the API substantially does not accumulate in the liver and is substantially not broken down by the liver before it has reached and had a therapeutic effect on the pancreas or one or more tissues and / or cells of the pancreas.

[0315] Accumulation in the pancreas may mean an amount of the API (in active form) reaches the pancreas which is effective to treat or mitigate a disease of the pancreas. Thus, accumulation in the pancreas may preferably be defined or summarised simply as the prevention, mitigation or treatment of a disease or disorder of the pancreas (or of one or more tissues and / or one or more cells thereof; such as pancreatic cancer, especially PDAC, or diabetes, such as by targeting beta cells) by the API. As used herein, the term disease or disorder of the pancreas may encompass diseases and disorders not traditionally categorised as being of the pancreas, but in which the pancreas nevertheless plays a role and / or in which targeting one or more tissues and / or cells of the pancreas may benefit; such, for example, as the targeting of beta cells in the prevention or treatment of diabetes.

[0316] Accumulation in the pancreas may suitably be measured by well-known techniques, such as by positron emission tomography (PET) of radio-labelled API; or optical imaging (e.g., IVIS imaging, as in Example 2 below) of fluorescently labelled API. These techniques may typically be non-invasive (especially, whole-body PET or whole-body IVIS). As such, accumulation in the pancreas may mean that at least about 30, 40, 50, 60 or 70 % of total API that is administered (such as administered orally or by injection) to the patient (or, total API that enters the bloodstream of the patient) enters the pancreas; which may be quantified by PET of radio-labelled API or optical imaging (e.g., IVIS) of fluorescently labelled API.

[0317] Thus, promoting the accumulation of the API in the pancreas may be or comprise effecting entry to the pancreas of at least about 30, 40, 50, 60 or 70 % of total API administered to the patient.

[0318] Where appropriate (e.g., if the API is administered orally), promoting the accumulation of the API in the pancreas may be or comprise effecting entry to the pancreas of at least about 30, 40, 50, 60 or 70 % of total API that enters the bloodstream of the patient.

[0319] Promoting the accumulation of the API in the pancreas may mean that fluorescence of fluorescently tagged API may be visible in the pancreas within about 24, 48 or 72 hours of administering the API, while such fluorescence is substantially not visible in the liver during the same time period following administration of the API.

[0320] Especially, promoting the accumulation of the API in the pancreas may mean that more (such as about 2x, 3x, 4x or 5x as much) API enters the pancreas, than enters the liver. Where no API enters the liver, most API may enter the pancreas.

[0321] The spleen; and promoting accumulation of an API (such as miRNA) therein

[0322] The spleen has a similar structure to a lymph node. The spleen functions primarily to filter the blood, including removing damaged and / or aged red blood cells. This role makes the spleen conducive to therapeutic strategies involving erythrocyte decoration or hitchhiking. Monocyte precursors stored in the spleen may act as Trojan horses for shaping immune responses and / or delivering drugs to tumours (or other dysregulated organs, tissues or cells). Targeted delivery of APIs to the spleen therefore offers the potential for immunomodulation, involving the treatment of various immune- related conditions, including different cancer types, infectious diseases, and autoimmune disorders.

[0323] The anatomy of the spleen is such as to present little or no sieving by endothelia. Thus, the amount of API that will enter into the spleen parenchyma is typically highly dependent on the amount of API capable of reaching the spleen via the blood. Although freshly ingested or injected API may in some instances reach the spleen before it reaches the liver, the API may quickly be metabolised by the liver, leading to a reduced steady state concentration developed over time in the blood and a reduced availability of the API to the spleen.

[0324] Compositions and methods in accordance with the present disclosure may be for promoting the accumulation of an API (especially, miRNA) in the spleen.

[0325] As used herein, the term accumulation in the spleen may mean accumulation in the spleen of an API for therapy of a splenic tissue and / or cell; such, in particular, as an miRNA for therapy of splenic cancer. The accumulation in the spleen may be or involve build-up of the API in one or more tissues of the spleen and treatment of a disease of the one or more tissues of the spleen by the API.

[0326] Accumulation in the spleen may mean that more than about 10, 20, 30 or 50 % of total API that enters the bloodstream is available (especially, available in active form, i.e., before it is metabolised, especially metabolised by the liver) for the treatment of one or more splenic tissues and / or cells.

[0327] Accumulation in the spleen may mean that the API substantially does not accumulate in the liver and is substantially not broken down by the liver before it has reached and had a therapeutic effect on the spleen or one or more tissues and / or cells of the spleen.

[0328] Accumulation in the spleen may mean an amount of the API (in active form) reaches the spleen which is effective to treat or mitigate a disease of the spleen. Thus, accumulation in the spleen may preferably be defined or summarised simply as the prevention, mitigation or treatment of a disease or disorder of the spleen (or of one or more tissues and / or one or more cells thereof; such as cancer of the spleen) by the API.

[0329] Accumulation in the spleen may suitably be measured by well-known techniques, such as by positron emission tomography (PET) of radio-labelled API; or optical imaging (e.g., IVIS imaging, as in Example 2 below) of fluorescently labelled API. These techniques may typically be non-invasive (especially, whole-body PET or whole-body IVIS).

[0330] As such, accumulation in the spleen may mean that at least about 30, 40, 50, 60 or 70 % of total API that is administered (such as administered orally or by injection) to the patient (or, total API that enters the bloodstream of the patient) enters the spleen; which may be quantified by PET of radio-labelled API or optical imaging (e.g., IVIS) of fluorescently labelled API.

[0331] Thus, promoting the accumulation of the API in the spleen may be or comprise effecting entry to the spleen of at least about 30, 40, 50, 60 or 70 % of total API administered to the patient.

[0332] Where appropriate (e.g., if the API is administered orally), promoting the accumulation of the API in the spleen may be or comprise effecting entry to the spleen of at least about 30, 40, 50, 60 or 70 % of total API that enters the bloodstream of the patient.

[0333] Promoting the accumulation of the API in the spleen may mean that fluorescence of fluorescently tagged API may be visible in the spleen within about 24, 48 or 72 hours of administering the API, while such fluorescence is substantially not visible in the liver during the same time period following administration of the API.

[0334] Especially, promoting the accumulation of the API in the spleen may mean that more (such as about 2x, 3x, 4x or 5x as much) API enters the spleen, than enters the liver. Where no API enters the liver, most API may enter the spleen. In accordance with the present disclosure, accumulation (as the term is defined herein) of an API in both the pancreas and spleen may be promoted by a composition defined herein. Accumulation of an API in both the pancreas and spleen, but not in the liver, may be promoted by a composition defined herein.

[0335] Accumulation in the pancreas and spleen may mean that an amount of total API administered to the patient, such as an amount consisting of at least about 30, 40, 50, 60 or 70 % of total API administered to the patient, enters the pancreas and / or the spleen. Preferably, the amount is distributed across both the pancreas and the spleen (for example, if the amount is at least about 50 % of total API administered to the patient, then at least about 20 % of total API administered to the patient may enter the spleen, while at least about 23 % of total API administered to the patient may enter the pancreas); which may be quantified by PET of radio-labelled API or optical imaging of fluorescently labelled API.

[0336] Thus, promoting the accumulation of the API in the pancreas and the spleen may be or comprise effecting the distribution of an amount of at least about 30, 40, 50, 60 or 70 % of total API administered to the patient, to a group of organs comprising or consisting of the pancreas and spleen.

[0337] Where appropriate (e.g., if the API is administered orally), promoting the accumulation of the API in the pancreas and the spleen may be or comprise effecting the distribution of an amount of at least about 30, 40, 50, 60 or 70 % of total API that enters the bloodstream of the patient, to a group of organs comprising or consisting of the pancreas and spleen.

[0338] Promoting the accumulation of the API in the pancreas and spleen may mean that fluorescence of fluorescently tagged API may be visible in the pancreas and spleen within about 24, 48 or 72 hours of administering the API, while such fluorescence is substantially not visible in the liver during the same time period following administration of the API. Especially, promoting the accumulation of the API in the pancreas and spleen may mean that more (such as about 2x, 3x, 4x or 5x as much) API enters the group of organs consisting of the pancreas and spleen, than enters the liver. Where no API enters the liver, most API may enter the spleen and / or pancreas.

[0339] Administration of compositions according to the disclosure

[0340] A composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) an active pharmaceutical ingredient (API; especially, miRNA), may be administered in a prophylactically effective amount to a human subject, wherein the subject is in need thereof; for example, identified by a physician or other healthcare practitioner as being in need thereof. Meanwhile, mitigating or treating a disease or disorder in a human subject may comprise administering a therapeutically effective amount of the composition to a human subject in need thereof.

[0341] Dosage amounts of the composition may be varied so as to obtain an amount of the API which is effective to achieve the desired prophylactic and / or therapeutic response for a given subject, without being toxic to the subject. A suitable dosage amount of the composition may be the amount of the composition which is the lowest dosage amount effective for the API to produce a therapeutic and / or prophylactic effect.

[0342] The selected dosage amount, form and regime will each depend upon a variety of factors. Such factors may include, for example, the activity of the API, the route of administration, the time of administration, the rate of excretion or metabolism of the API, the rate and extent of absorption, the duration of the treatment, the presence of other drugs, compounds and / or materials used in combination with the API, the age, sex, weight, condition, general health and prior medical history of the subject being treated, and other such factors well known in the medical arts.

[0343] The composition may be administered by intramuscular or intravenous injection (encompassing transdermal delivery via a patch), orally (encompassing sublingual administration), intranasally, dermally, or by any other suitable route; especially by injection or orally. Preferably, the composition may be administered by injection, such, for example, as intravenous or intramuscular injection. Optionally when the composition is administered by injection, the subject is monitored for symptoms or signs of a hypersensitivity response, such, for example, as a vaccine-associated hypersensitivity response.

[0344] Also preferably, the composition may be administered orally or intranasally. Compositions suitable for oral administration may be presented as discrete dosage forms, especially liquids or aerosol sprays each containing a predetermined amount of the composition. Such dosage forms may be prepared by any of the well-known methods of pharmacy.

[0345] Thus, it is especially preferred that the composition is for use in a method of preventing or treating a disease or disorder in a human subject, wherein the method comprises administering the composition to the human subject by injection, orally or intranasally (especially, by injection or orally). A murine model of administration by injection is set out, for instance, in the Examples.

[0346] The composition may be combined in an intimate admixture with a pharmaceutical carrier, according to conventional pharmaceutical compounding techniques. The carrier may take a wide variety of forms depending on the form of preparation desired for administration. Any of the usual pharmaceutical media may be employed as carriers, such as, for example, one or more of water, oils, and alcohols (encompassing glycols). The forms in which the disclosed compositions may be incorporated for administration, especially when formulated for administration by injection, orally or intranasally, may include aqueous solutions in saline. The composition may further include one or more pharmaceutically acceptable additives and excipients, such as one or more of the following: detackifiers, anti-foaming agents, buffering agents, polymers, antioxidants, chelating agents, viscomodulators, tonicifiers, odorants, opacifiers, suspending agents, fillers, plasticizers, flavouring agents, preservatives, colouring agents, diluents, binders, disintegrating agents and mixtures thereof. The prevention or attenuation of the action of microorganisms may be brought about by the inclusion of various antibacterial and antifungal agents; such as one or more of the following: parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like.

[0347] The composition disclosed herein may be provided in a sterile solution, by incorporating the composition in the required amount in an appropriate solvent (with various other ingredients, where appropriate) by any of the well-known methods of pharmacy. The composition disclosed herein may be provided in a sterile dispersion, by incorporating the composition in the required amount in an appropriate sterile vehicle (with various other ingredients, where appropriate). The composition disclosed herein may be provided as a sterile powder (e.g., for the subsequent preparation of sterile injectable solutions), such as by vacuum-drying and freeze-drying (lyophilisation) techniques which yield a powder of the composition.

[0348] Stabilising effects of compositions according to the disclosure

[0349] Optionally, a silicon- and lipid-containing delivery vehicle in accordance with the present disclosure (e.g. 3C-SiAEN-RE0.1, 3C-SiLB-RE0.1, 2C-SiAEN-Tyr-RE0.1, 2C-SiLB-RE0.1, 2C-LBSi-Tyr-RE0.1 or 2C-LBSi-Ch-OSA0.3-Tyr-Gly-RE0.1) is prepared, stored, and delivered to a clinic, then complexed with an API, such as mRNA or miRNA (especially, miRNA), before it is administered to a patient. Optionally, the delivery vehicle is stored separately from the API (such as mRNA or miRNA, especially miRNA), until close to the time when it is administered to a patient. Since the delivery vehicle does not contain the reactive API while it is being stored, there is no need for storage at especially cold temperatures such as below 4 °C in order specifically to stabilise the API. This is a different approach to conventional lipid nanoparticle delivery vehicles, which are usually stored already encapsulating an API and which typically need to be stored at temperatures matching the API, such as at ultralow temperatures if the API is prone to degradation.

[0350] Moreover, the doped Si-containing delivery vehicle disclosed herein may have a stabilising effect on a reactive API, such as a nucleic acid (especially, mRNA or miRNA, most especially miRNA) when complexed with it. As a result, the delivery vehicle of the disclosure may optionally be stored already complexed to the API. This may be possible at not especially cold temperatures. For example, the composition comprising the API could be stored at temperatures of about 0 °C or above, especially of about 3-5 °C, i.e. the temperature of a typical commercially available refrigerator; or at about 15-30 °C, i.e. typical room temperature.

[0351] The Si-containing delivery vehicle may also stabilise APIs while circulating in the body. Additionally or alternatively, it can ensure efficient API uptake by cells. Additionally or alternatively, it can stabilise the API in the cytoplasm of a cell. For example, this may be advantageous where the API is mRNA which is preferably delivered safely to ribosomes in the cell cytoplasm for translation.

[0352] Thus, a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon may be stored, either before or after combination with (iii) the API. As shown in the Examples hereinbelow, in contrast to conventional liposomal delivery vehicles wherein an API must be added at the same time as the liposome is formed (since the API will be encapsulated in the liposomal cavity), it has been found that an API delivery vehicle comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, may be formed first, then (iii) the API may be added.

[0353] Optionally, the composition (with or without the API) is stored at temperatures in a range of from about 3 °C to about 50 °C, especially in a range of from about 15 °C to about 25 °C. Thus, the composition may be stored at standard refrigerator temperature (about 5 °C), room temperature (about 20°C), or higher temperatures (for example, about 30, about 40 or about 50 °C). The composition has been found to be storage stable at such temperatures, for example not degrading for a period of at least about 1, 6 or 12 months. This is conducive not only to efficient storage but also to efficient transport and distribution of the composition.

[0354] Optionally, therefore, the composition (with or without the API) is stored at a temperature in a range of about 0 °C or above, especially about 4 °C or above, for a period of at least about 1 week (optionally, up to about 6 months, especially up to about 1 year) prior to administering the composition to the subject. In certain preferred embodiments, the composition is stored without the API. Shortly before administration to the subject, such as not more than about 3 weeks, about 2 weeks or about 1 week, especially not more than about 2 days, more especially not more than about 1 day before administration to the subject, they will be combined with the API (for example, via steps such as those described in the Examples hereinbelow). Advantageously, the particles and lipid may be storage stable for an extended period of time, especially a period of several months, such as about 6 months or about 12 months; they may be stored in this way without appreciable degradation. When the API is a reactive API, especially mRNA or miRNA, it may advantageously be synthesised in situ or synthesised off-site and delivered to a clinical setting at the last possible moment, then combined with the particles and lipid(s).

[0355] Nonetheless, it has also been found that an API can be stabilised by the particles comprising hydrolysable doped silicon, including that the API can be stabilised during storage. Thus, in some embodiments, the particles can increase the storage stability of the API, especially where the API is or comprises nucleic acid such as mRNA or miRNA. In some embodiments, the particles can increase the stability at 25 °C of the API, especially where the API is or comprises nucleic acid such as mRNA or miRNA.

[0356] Thus, optionally, the composition being stored comprises all of the following components: (i) the one or more lipids, (ii) the particles comprising hydrolysable silicon and (iii) the API. If so, the composition, once formulated, may optionally be stored for only a short period, by which may be meant a period of up to about 3 weeks, about 2 weeks or about 1 week prior to administration to the subject.

[0357] The particles may increase the stability of the API during circulation in vivo, especially where the API is or comprises nucleic acid such as mRNA or miRNA. Additionally or alternatively, the particles may protect the API from degradation, especially enzymatic degradation, especially where the API is or comprises nucleic acid such as mRNA or miRNA. Thus, the compositions disclosed herein may mitigate or address the problem of how to ensure APIs reach cells once they have been administered to a patient, including how to stabilise APIs while they are circulating in the body.

[0358] Targeting effects of compositions according to the disclosure

[0359] Meanwhile, the disclosed composition comprising hydrolysable doped silicon may mitigate or meet the need for organ, tissue and / or cell targeting, so that an API (especially, miRNA) can be delivered to the correct cells.

[0360] Additionally or alternatively, once a target organ, tissue and / or cell is reached, the disclosed composition may mitigate or solve the challenge of how to ensure efficient API uptake by the organ, tissue and / or cell. For example, the disclosed composition may assist trafficking of the API from the exterior of a cell into the cytoplasm.

[0361] Following uptake of the API by a cell, the disclosed composition may mitigate or solve the problem of how to prevent the API degrading too quickly in the cytoplasm. It is thought that the particles can increase the stability of the API in the cytoplasm of a cell, especially stability against enzymatic degradation, especially where the API is or comprises nucleic acid such as mRNA or miRNA, especially miRNA.

[0362] In a particularly preferred scenario, the composition may mitigate or solve the problem of how to prevent the API being sequestered and metabolised by the liver. Surprisingly, as the Examples below show, the composition may enhance uptake of an API by the pancreas and / or spleen, instead of by the liver.

[0363] Without wishing to be bound by theory, it is proposed that the preferential uptake of the API by the pancreas and / or spleen may be due to the surface characteristics, such as roughness and / or surface charge, of the particles associated with lipid and API. It is thought that the particles’ “rough” surface may play a role in their bypassing the liver in favour of the pancreas and / or spleen. Meanwhile, secondary accumulation of the particles may occur in tissues having exceptionally high blood perfusion, such as the gonadal tissues exemplified in the Examples below. Preferred action of compositions according to the disclosure

[0364] As illustrated by the Examples below, compositions according to the disclosure may “bypass” the liver and accumulate in the pancreas and / or the spleen. The preferential transfection of pancreatic and splenic cells in vivo in mice with, variously, mRNA and siRNA, shows that compositions according to the disclosure can not only reach the pancreas and / or spleen in sufficient amounts, but also deliver functional API to the cells thereof, so that it is taken up and acts on the cells (here, translated, with consequent expression of observable bioluminescence or fluorescence). Importantly, “bypassing” the liver means that API is available to the pancreas and / or spleen which might otherwise not be.

[0365] The present disclosure therefore especially provides the compositions described herein for the prevention, treatment or mitigation of a disease or disorder of the pancreas (especially cancer, most especially PDAC) and / or of the spleen.

[0366] Preferred compositions comprise (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) the API, wherein the one or more lipids consist essentially of or comprise one or more cationic lipids (e.g. DOTAP); one or more phospholipids (e.g. DOPE); and / or one or more polyethylene glycol (PEG) lipids (e.g. DSPE-PEG2000); and the particles comprising hydrolysable silicon are porous particles (optionally, doped, e.g. boron-doped; optionally, surface treated with one or more alcohols, e.g. methanol) comprising at least 50 % w / w elemental silicon and having a mean diameter of about 1 to about 100 nm (e.g., 30 nm). The composition preferably further comprises one or more amino acids (especially tyrosine, arginine and / or glycine). The composition preferably further comprises trehalose. The composition may further comprise ortho silicic acid (OSA) or stabilised OSA (e.g. choline-stabilized ortho silicic acid, known as “Ch-OSA”); without wishing to be bound by theory, it is thought that this may facilitate lipid-silicon binding, as described herein.

[0367] The ratio by weight of total lipid (i) to the particles (ii) may preferably be about 40: 1 to about 1 : 1, especially about 20: 1 to about 1 : 1; such, for example, as a ratio by weight of about 16: 1, when the components are assembled for manufacture of a delivery system, i.e. before any further processing is carried out (such further processing may be, for example, the processing under ""Extrusion" of the Examples hereinbelow).

[0368] Preferably, the API is or comprises RNA, especially single-stranded RNA such as mRNA or miRNA, especially miRNA.

[0369] The ratio by weight of total lipid (e.g., where the lipids consist of DOPE and DOTAP, the weight of total lipid is the weight of the DOPE and the weight of the DOTAP, combined) to total API may preferably be about 5: 1 to about 100: 1, especially about 5: 1 to about 50: 1. Examples include a ratio by weight of total lipid to API of about 12: 1, about 24: 1 and about 48: 1.

[0370] Examples

[0371] Example 1

[0372] Example 1 provides a protocol for the preparation of silicon-based delivery systems loaded with an active pharmaceutical ingredient (API), using formulations 3C- SiAEN-REO. l and 3C-SiLB-RE0.1 as representative delivery vehicles; and mRNA as a representative API.

[0373] An analogous protocol may be followed to prepare other Si-containing delivery platforms (such as those containing different lipids, different amino acids, etc.) and to complex them with mRNA or with another API.

[0374] Delivery vehicles 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1

[0375] Delivery vehicles 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1 were prepared having the following formulations. To summarise, 3C-SiAEN-RE0.1 contains undoped Si nanoparticles, whereas 3C-SiLB-RE0.1 contains Si nanoparticles doped with circa 5xl018boron atoms / cm3. (This applies throughout the present specification where “SiAEN” (referring to the undoped version) or “SiLB” (referring to the doped version) is used.)

[0376] Table 1 .

[0377] Additional details of the above-disclosed components of 3C-SiAEN-RE0.1 and 3C- SiLB-REO. l are as follows. a) Si (“SiNPs ”) : as porous silicon particles of 30 nm average diameter (other suitable average diameters include, for example, 10 nm, 30 nm, 50 nm or 100 nm). Boron doped for 3C-SiLB-RE0.1. Activated by exposure to methanol followed by slow evaporation, producing (dry, solid) powder, which is activated SiNPs. (Analogous particles were provided for use in the other compositions of the present Examples.) b) Trehalose (“THR”): as solid powder. c) Glycine (“GLY”) : as solid powder. d) SiNPs + GLY + THR solution: as a brownish suspension, obtained via sonication for 60 minutes of a suspension in 50 ml nuclease-free water of: 50 mg activated SiNPs; 50 mg THR; and 25 mg GLY. e) DOTAP-C1 Solution: as a solution obtained by mixing 50 mg DOTAP in 10 ml of methanol, followed by sonication at 40 °C for 30 minutes, until fully dissolved. f) DOPE Solution: as a solution, obtained by mixing 50 mg DOPE in 10 ml of methanol, followed by sonication at 40 °C for 30 minutes, until fully dissolved. g) mPEG2000-DSPE Solution: as a solution, obtained by mixing 40 mg of mPEG2000-DSPE in 8 ml of methanol, followed by sonication at 40 °C for 30 minutes, until fully dissolved. h) Sialic acid: as a solution in methanol.

[0378] Method for preparing 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1 delivery vehicles

[0379] Lipid film preparation a) Mix all lipids (DOPE, DOTAP and DSPE-PEG2000) in a glass round bottomed flask. b) Evaporate solvent with rotary evaporator in water bath at 40 °C.

[0380] Rehydration of the film a) Add 1 ml of the brownish suspension of SiNPs (3C-SiLB-RE0.1 : doped;3C- SiAEN-REO.l: undoped) + GLY + THR (Img / ml SiNPs; 0.5 mg / ml GLY; Img / ml THR) and 0.4 ml sialic acid to the lipid film, along with 8.6 ml nuclease free water (total volume of the brownish suspension and nuclease free water together is thus 10 ml). b) Cover the flask with parafilm, then agitate the flask in a water bath at 60 °C for 10 minutes, thus rehydrating the lipid film by means of the 10ml of liquid. c) Leave resultant suspension to rest at room temperature for a few hours before storing at 4 °C. Extrusion

[0381] Pass the suspension obtained in step (c) of “Rehydration of the film” through a polycarbonate membrane filter having 0.4 pm and 0.1 pm pore sizes. Pass the suspension 10 times, at 60 °C, through each pore size.

[0382] The resultant products are 3C-SiAEN-RE0.1 where undoped SiNPs are used and 3C-SiLB-RE0.1 where doped SiNPs are used.

[0383] Method for preparation of complexes of 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1 delivery vehicles with mRNA

[0384] An exemplary mRNA for complexation with the silicon-containing delivery vehicles is DasherGFP mRNA (commercially available from Aldevron) coding for green fluorescent protein (GFP).

[0385] For preparation of 3C-SiAEN-RE0.1-mRNA complexes, a suspension containing 3C-SiAEN-RE0.1 having a total lipid fraction (DOPE, DOTAP and DSPE- PEG2000, taken together) of 120 pg is mixed with 10 pg mRNA. This achieves a ratio by weight of total lipids to mRNA of 12: 1.

[0386] For preparation of 3C-SiLB-RE0.1-mRNA complexes, a suspension containing 3C-SiLB-RE0.1 having a total lipid fraction (DOPE, DOTAP and DSPE-PEG2000, taken together) of 120 pg is mixed with 10 pg mRNA. This achieves a ratio by weight of total lipids to mRNA of 12: 1.

[0387] It will be appreciated that different ratios may be used, as required.

[0388] For complexation, the samples are mixed thoroughly by gently pipetting and are incubated at room temperature for 60 min to allow for complexation to complete.

[0389] Following incubation and thus complexation of the mRNA with the Si- containing delivery platforms, the complexed samples are typically stored at 4 °C prior to use in subsequent experiments. Additional delivery vehicles

[0390] Other delivery vehicles that may be prepared in an analogous procedure to that set out above for 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1, are detailed below.

[0391] 2C-SiAEN-RE0.1 and 2C-SiLB-RE0.1 are used in Example 2 below:

[0392] 2C-SiAEN-Tyr-RE0.1 incorporates tyrosine:

[0393] 2C-SiLB-RE0.1 replaces undoped SiNPs in 2C-SiAEN-RE0.1 with doped SiNPs (circa 5xl018boron atoms / cm3):

[0394] 2C-LBSi-Tyr-RE0.1 incorporates tyrosine:

[0395] Optionally in 2C-LBSi-Tyr-RE0.1, DOPE-CF (CAS No. : 384832-91-1; DOPE labelled with a fluorescein moiety) may be added, to form 2C-LBSi-Tyr-RE0.1 tagged with DOPE-CF (for imaging purposes):

[0396] 2C-AESi-Tyr-RE0.1 also incorporates tyrosine:

[0397] Other formulations include:

[0398] *bPEI: branched polyethyleneimine; included in the “lipids” fraction for preparation purposes, according to the protocol of Example 1; although bPEI may not strictly be considered a lipid, but rather a transfection reagent to assist with endocytosis

[0399] (undecyloxy)hexyl]amino}octanoate. CAS No. : 2089251-47-6.

[0400] ** ALC-0315 (DTA-PEG2000): 2-hexyl-decanoic acid, l,l'-[[(4- hydroxybutyl)imino]di-6,l-hexanediyl] ester, CAS No. 2036272-55-4; included in the “lipids” fraction for preparation purposes, according to the protocol of Example 1; although it may not always be considered a strict lipid.

[0401] * Choline-stabilized orthosilicic acid. Included in the “lipids” fraction for preparation purposes, according to the protocol of Example 1; although it may not strictly be considered a lipid, but rather an agent to assist with lipid-Si binding.

[0402] Example 2 - Checking transfection efficacy in a human pancreatic adenocarcinoma cell line model

[0403] This study was to validate the transfection efficiency of 2C-SiAEN-RE0.1 and 2C-SiLB-RE0.1 delivery vehicles in vitro, in a human pancreatic adenocarcinoma cell model. The human cell line PSN1, derived from a pancreatic adenocarcinoma, was used.

[0404] 2C-SiAEN-RE0.1 and 2C-SiLB-RE0.1, the formulations of which are set out in Example 1 above, were formulated by following an analogous protocol to that set out in Example 1 for 3C-SiAEN-RE0.1 and 3C-SiLB-RE0.1.

[0405] 2C-SiAEN-RE0.1 and 2C-SiLB-RE0.1 were each complexed with commercially available DasherGFP mRNA (Aldevron) coding for green fluorescent protein (GFP), at five different ratios by weight of total lipids to mRNA: 4: 1 (Rl), 7.2: 1 (R2), 12: 1 (R3), 24: 1 (R4) and 48: 1 (R5)..

[0406] Transfection efficiency was compared to that of commercial transfecting reagents, namely Dharmafect and Lipofectamine, to provide a positive control. Dharmafect and Lipofectamine results were obtained at 5, 2 and 0.5 pl concentrations of each reagent, to provide a frame of reference for the tests using 2C-SiAEN-RE0.1 and 2C-SiLB-RE0.1.

[0407] Cells were transfected in a routine manner (seeded at 10,000 cells / well). GFP expression was assessed after 24h and 48h post transfection using a commercially available FLUOstar Omega (BNG LABTECH). Adequate transfection efficiency was confirmed at each of the tested ratios, by adequate GFP expression at 24h and 48h, in comparison to Dharmafect and Lipofectamine.

[0408] Example 3 - Murine model

[0409] This Example studies the targeting of the pancreas and spleen by delivery vehicles in the scope of the present disclosure when loaded with reporter mRNA; and their “bypassing” of the liver.

[0410] Six one-month old albino CD1 mice (obtainable from Charles River Laboratories of Keele and Alderley Park, UK) were injected intraperitoneally (IP) with 3C-SiAEN-RE0.1 complexed with mRNA, or 3C-SiLB-RE0.1 complexed with mRNA (a single injection per mouse per time point, of 1000 pmol in a volume of 50 pl). The mice were then studied at time points of 6 hours, 24 hours and 5 days post injection.

[0411] A firefly luciferase-coding mRNA reporter having 2315 bases was used, for the reason that its expression could easily be visually monitored. As well as other mRNAs, the successful delivery of the mRNA-fireflyLuc reporter is expected to provide evidence that miRNA delivery would occur in a similar manner.

[0412] As in the exemplary protocol set out in Example 1, the ratio by weight of the total lipid fraction of 3C-SiAEN-RE0.1 to mRNA was 12:1; and the ratio by weight of the total lipid fraction of 3C-SiLB-RE0.1 to mRNA was also 12: 1.

[0413] Mice were anaesthetised at the 6-hour and 24-hour time points and were imaged using in vivo fluorescence imaging (IVIS) using a Hamamatsu 1394 ORCA II Deep Cooling BTA 1024 IVIS imaging device (obtainable from Hamamatsu Photonics of Shizuoka, Japan). Region of interest (ROI) were drawn for each mouse to capture its whole body, and total flux (p / s) was reported.

[0414] Figure la shows fluorescence at 6 hours, while Figure lb shows fluorescence at 24 hours. The site of intraperitoneal injection (1001) shows fluorescence, as do, notably, the pancreas (1003) and spleen (1004); and, to some extent, white adipose tissue of the gonads (1002) (the latter of which is highly vascularised). Fluorescence of the liver is unexpectedly low (it is minimal to zero).

[0415] After 5 days, mice were culled humanely and their organs were washed and resuspended in luciferin-containing medium. IVIS imaging of the organs is shown in Figure 2. Again, the pancreas and spleen show fluorescence; as does, to some extent, white adipose tissue of the gonads, as shown by dotted circles around the relevant areas (the original colour photograph from which Figure 2 is prepared shows bright fluorescence in the areas now circled in dotted black lines). The fluorescence of the white adipose tissue is thought to be due to the exceptionally high blood perfusion in such tissues. The fluorescence of the pancreas and spleen is unexpected. Fluorescence in the liver might have been expected (in view of its typical role in sequestering and breaking down API, as described herein), but was not observed. Surprisingly, a silicon- containing delivery platform as tested appears to assist in targeting the investigative API to pancreatic tissue having dense fibrotic stroma and to the related tissue of the spleen; while “bypassing” the liver.

[0416] Example 4 - Further murine model

[0417] Following an analogous procedure to Example 3, a second murine model is established, except that the mRNA of Example 3 is replaced by an investigative siRNA, namely fluorescent Cy5.5-siRNA, commercially available from Sigma Aldrich; and that delivery with 3C-SiAEN-RE0.1, but not with 3C-SiLB-RE0.1, was tested. As in Example 3, fluorescence is detected using in vivo fluorescence imaging (IVIS) with a Hamamatsu 1394 ORCA II Deep Cooling BTA 1024 IVIS imaging device (obtainable from Hamamatsu Photonics of Shizuoka, Japan).

[0418] To follow the time progression in more detail than in Example 3, at each of the time points O.lhr, Ihr, 2hr, 4hr, 6hr and 24hr following administration of 3C-SiAEN- RE0.1 loaded with Cy5.5-siRNA by IP injection, a mouse was randomly selected for whole-body live animal imaging using the IVIS system. The results are shown in Figures 3a, 3c, 3e, 3g and 3i.

[0419] The additional mouse on the left of Figure 3a is untreated (control). Fluorescence is observed in the organs of the gastrointestinal track (stomach, intestine) in both the untreated (control) mouse and in the treated mouse and is therefore assigned to background fluorescence (such as that emanating from feed).

[0420] After whole body imaging, the mouse was culled humanely and the following organs were resected: Liver, Kidney, Intestine, Stomach, Spleen, Injection site (Fat pad), Bladder, Lungs, Heart, Blood. The organs were washed in PBS, arranged on a petri dish and imaged to quantify the Cy5.5-siRNA signal in the tissues. The results are shown in Figures 3b, 3d, 3f, 3h and 31.

[0421] A small section of each tissue (weighing approximately 30mg) was siphoned to a SPEX Sample Prep tube and frozen in liquid nitrogen. The samples were homogenised and the amount of siRNA in each was quantified using SL-qPCR. The results are shown graphically in Figure 4 and are tabulated in Tables 15 and 16:

[0422] Quantified fluorescence as Avg Radiant Efficiency of each organ, following IP injection with 1000 pmol of Cy5.5-siRNA (loaded to Si-containing delivery vehicle), at timepoints 0. Ih, Ih, 2h, 4h, 6h, and 24h. The spleen vs liver accumulation was assessed, as shown in the final row. Increased accumulation in the spleen compared to the liver was observed at an early time point. Table showing Ct Value and amount of siRNA in the liver at T=0.1, 1, 2, 4 and

[0423] 6 hours after IP injection. Measured using SL-qPCR.

[0424] Thus, Figure 4 shows a line graph of the fluorescence (as average radiant efficiency) quantified in each organ at timepoints O. lh, Ih, 2h, 4h, 6h, and 24h. A larger group of organs fluoresced at the earlier time timepoints (O.lh to 2h). This suggests ready dispersion of the siRNA complex from the site of IP injection around the peritoneal cavity readily.

[0425] Surprisingly, preferential accumulation was observed in the spleen compared to the liver at earlier timepoints, when the ratio of accumulation in the spleen compared to the liver was approximately 2: 1, even without normalisation by weight. This suggests that 3C-SiAEN-RE0.1 loaded with Cy5.5-siRNA “bypassed” the liver and was able to target the more difficult-to-reach spleen and, it is inferred (by reference to the physiological principles described herein and by analogy with Example 3), the pancreas, rather than the siRNA being sequestered by and broken down in the liver. Significant “build up” in the liver was not observed at any time point.

[0426] Apparent “accumulation” in the kidney and bladder, for example as seen in Figure 4, may be attributed to the size of these organs and the non-normalisation of the data by weight. The kidney, in particular, being larger than the other tested organs, has a disproportionate average radiant efficiency until normalised by weight.

[0427] Thus, Example 4 provides evidence for “bypassing” of the liver, in preference for the spleen and - by reference to the physiological principles described herein and by analogy with Example 3 - the pancreas, with 3C-SiAEN-RE0.1 loaded with Cy5.5- siRNA: an analogous formulation to Example 3 except replacing mRNA as the API, with siRNA. The levels of siRNA in other organs observed is in keeping with the expected passage of API following IP injection and background fluorescence of the digestive organs is as expected and as confirmed by the control mouse.

[0428] Example 5 - Further murine model

[0429] In Example 5, the murine model of Examples 3 and 4 was further developed.

[0430] An analogous protocol was followed to that of Examples 3 and 4, except that a larger cohort of mice was provided, the mice were c57BL / 6 mice aged 8 weeks (obtainable from Charles River); and different silicon-containing delivery platforms were used. The silicon-containing delivery platforms and mRNAs set out in Table 17 (see Example 1 for formulations) were used. 3 mice were provided per group. Dosing was via a single intravenous injection to the tail.

[0431] * The same firefly luciferase-coding mRNA reporter having 2315 bases as in Example 3.

[0432] **mRNA GFP-Cy5, commercially available from Stratech.

[0433] Results were obtained from each group 1-10 using IVIS (as in Examples 3 and 4) to observe bioluminescence (relevant to luc mRNA) or fluorescence (relevant to GFP-Cy5; the characteristic CY5 fluorescence Em peak is at 667 nm). For Group 1 (saline control group), no luminescence or fluorescence was observed. For Group 2 (naked mRNA control group), no significant luminescence or fluorescence was observed. A slight fluorescence was observed in the gastrointestinal tract and attributed to background fluorescence, similar to that observed in Example 4 above.

[0434] For Group 3 (empty Si-containing delivery vehicle control, having DOPE tagged with carboxylfluorescin: 2C-LBSi-Tyr-RE0.1 tagged with DOPE-CF), no luminescence or fluorescence was observed. It appears that the small volume of tagged DOPE may not have generated adequate fluorescence for IVIS imaging.

[0435] No CY5 fluorescence is observed for Group 4 (as expected). The bioluminescence results for Group 4 are shown in Figures 5a to 5c, respectively showing bioluminescence at 6, 24 and 48 h following IV injection. Figure 5a shows two images (A) and (B), as the signal was measured twice, the second time (B) without the mouse with the strongest signal (thought and confirmed to be masking the signal of the other two mice). Bioluminescence suggests a trend of preferential spleen and pancreas uptake, with some uptake by the liver but a greater, surprising uptake by the spleen and pancreas. Some bioluminescence is observed in the tail (site of injection).

[0436] Group 4 may be compared to Groups 8 to 10, which were analogous to Group 4 but with different components in the silicon-containing delivery vehicle.

[0437] No bioluminescence is observed for Group 5 (as expected). The CY5 fluorescence results for Group 5 are shown in Figures 6a to 6c, respectively showing CY5 fluorescence at 6, 24 and 48 h following IV injection. The sections of Figures 6a and 6b bordered by dotted black lines isolate the strongest fluorescence signals (by setting the lower limit of quantification to exclude low-level and background fluorescence). The trend is for the spleen and pancreas to show strong signals, which is especially clear when the lower limit of quantification is adjusted as described. Some fluorescence is observed in the tail (site of injection). No bioluminescence is observed for Group 6 (as expected). The CY5 fluorescence results for Group 6 are shown in Figures 7a to 7c, respectively showing CY5 fluorescence at 6, 24 and 48 h following IV injection. The sections of Figures 7a and 7b bordered by dotted black lines isolate the strongest fluorescence signals (by setting the lower limit of quantification to exclude low-level and background fluorescence). The trend is for the spleen and pancreas to show strong signals, which is especially clear when the lower limit of quantification is adjusted as described. Some fluorescence is observed in the tail (site of injection).

[0438] No bioluminescence is observed for Group 7 (as expected). The CY5 fluorescence results for Group 7 are shown in Figures 8a to 8c, respectively showing CY5 fluorescence at 6, 24 and 48 h following IV injection. The sections of Figures 8a and 8b bordered by dotted black lines isolate the strongest fluorescence signals (by setting the lower limit of quantification to exclude low-level and background fluorescence). As for group 6, the trend is for the spleen and pancreas to show strong signals, which is especially clear when the lower limit of quantification is adjusted as described. Some fluorescence is observed in the tail (site of injection).

[0439] No fluorescence is observed for Group 8 (as expected). The bioluminescence results for Group 8 are shown in Figures 9a to 9c, respectively showing bioluminescence at 6, 24 and 48 h following IV injection. Similarly to Figure 5a, Figure 9a shows three images (A) to (C), as the signal was measured three times to ensure mice with stronger signals were not masking the signal of other mice. The trend in bioluminescence suggests preferential spleen and pancreas uptake. Some bioluminescence is observed in the tail (site of injection).

[0440] No fluorescence is observed for Group 9 (as expected). The bioluminescence results for Group 9 are shown in Figures 10a to 10c, respectively showing bioluminescence at 6, 24 and 48 h following IV injection. Similarly to Figures 5 and 9a, Figures 10a and 10b show three images (A) to (C) and Figure 10c shows two images (A) and (B), as the signal was measured three (or two) times to ensure mice with stronger signals, and / or the signals emanating from the tails (site of injection) of the mice, were not masking other signals. The trend in bioluminescence suggests preferential spleen and pancreas uptake. Some bioluminescence is observed in the tail (site of injection).

[0441] No fluorescence is observed for Group 10 (as expected). The bioluminescence results for Group 10 are shown in Figures Ila to 11c, respectively showing bioluminescence at 6, 24 and 48 h following IV injection. Similarly to Figures 5, 9a, and lOa-lOc, Figure Ila shows three images (A) to (C) and Figures 11b and 11c shows two images (A) and (B), as the signal was measured three (or two) times to ensure mice with stronger signals, and / or the signals emanating from the tails (site of injection) of the mice, were not masking other signals. The trend in bioluminescence suggests preferential spleen and pancreas uptake.

[0442] Example 6 - Further murine model

[0443] Example 6 aims to further investigate the biodistribution results for the firefly luciferase-coding reporter mRNA delivered in accordance with Example 5, by comparing biodistribution in the spleen (hard to target) vs. the liver (difficult to “bypass”).

[0444] Following an analogous procedure to Example 5, mice were injected (IV, tail) with formulations as shown in Table 18 below.

[0445] 6 hours post-injection, mice were culled humanely and their spleens and livers were isolated and snap froze in liquid nitrogen. The spleens and livers’ luciferase activity was quantified using a Bright-Glo™ Luciferase Assay System, obtainable from Promega.

[0446] Comparison of the spleen to liver luciferase activity revealed statistically significant targeting of the spleen over the liver for at least test groups 4, 5 and 7, as shown in Figure 12. There appears to be a similar trend for group 6, but it is less significant. Representative results are shown in further detail in Figures 13a-c.

[0447] Overall, there is provided evidence of unexpectedly “bypassing” the liver and reaching the harder-to-target spleen, using various different compositions in the scope of the present disclosure which are loaded with RNA (luc mRNA).

[0448] Example 7 - miRNA delivery

[0449] Example 7 aims to investigate the delivery of miRNA using various different compositions in the scope of the present disclosure as the delivery vehicle.

[0450] The compositions used were: a. 2C-SiAEN-RE0.1 (for conciseness, labelled as SP1 below) b. 2C-SiAEN-Tyr-RE0.1 (SP2 below) c. 2C-SiLB-RE0.1 (SP3 below) d. 2C-LBSi-Tyr-RE0.1 (SP 4 below) e. 2C-LBSi-Ch-OSA0.3-Tyr-Gly-RE0.1 (SP 5 below)

[0451] These had the ingredients set out in Example 1 above and were prepared in accordance with the protocol established in Example 1.

[0452] Example 7a

[0453] A multi-well 2D culture of human pancreas adenocarcinoma (PA-TU-8988T) cells was established under standard conditions. Two different (proprietary) miRNAs (miRNAl and miRNA2) were tested.

[0454] Different amounts of each miRNA were tested: f. 5 nM g. 10 nM h. 20 nM i. 40 nM

[0455] Each miRNA amount was tested at two different ratios to the total lipid fraction of SPl to SP5: miRNA : total lipid fraction = 1 :24 miRNA : total lipid fraction = 1 :48

[0456] The end volume of liquid supplied to each well was made up to 180 pL across all wells.

[0457] After overnight incubation, cells were detached and re-seeded in low-adhesion plates. The formation and growth of tumour spheroids (clumps of tumour cells) was assessed over 5 days.

[0458] The inhibition by miRNA of tumour development was significant after only 3 to 5 days, as shown in Figure 14 (3 days) and Figure 15 (5 days), especially for miRNAl. Figures 13 and 14 exemplify 40 nM miRNA at miRNA : total lipid fraction = 1 :48, for each of SP1 to SP5 and for each of miRNAl and miRNA2, compared to untreated cells and control (no miRNA) cells.

[0459] Example 7b

[0460] An analogous procedure was followed to that of Example 7a, except that instead of being detached and re-seeded in low-adhesion plates, the cells remained in their original culture for 5 days. At the 5-day point, WST-1 assay (cell viability assay; kit obtainable from Sigma Aldrich) was used to assess cell viability. (The WST-1 assay protocol is based on the cleavage of the tetrazolium salt WST-1 to formazan by cellular mitochondrial dehydrogenases.) The results are shown in Figures 16a-e. As for Figures 14 and 15, Figures 16a-e exemplify 40 nM miRNA at miRNA : total lipid fraction = 1 :48, for each of SP1 to SP5 and for each of miRNA 1 and miRNA2, compared to untreated cells and control (no miRNA) cells. There is generally a large drop in absorbance for both miRNAl and miRNA2, which is evidence of successful transfection of the human pancreas adenocarcinoma (PA-TU-8988T) cells leading to reduced viability and thus an antitumour effect (the only exception being miRNAl for SP4, which may possibly be due to experimental failure; in any case, the general trend is compelling).

[0461] Example 7 thus supports the use of compositions encompassed by the disclosure for delivering miRNA to treat cancer, especially cancer of the pancreas.

[0462] Example 8 - Further murine model

[0463] In Example 8, the murine model of Example 5 was further developed.

[0464] An analogous protocol was followed to that of Example 5, except that different silicon-containing delivery platforms were used. The same controls (saline and naked mRNA) were used as in Example 5 (see Table 17 for details).

[0465] The silicon-containing delivery platforms and mRNAs set out in Table 19 were used. See Tables 20-24 for their formulations, which were prepared in the same way as the formulations detailed in Example 1.

[0466] 3 mice were provided per group. Dosing was via a single intravenous injection to the tail, as in Example 5.

[0467] * The same firefly luciferase-coding mRNA reporter having 2315 bases as in Examples 3 and 5.

[0468] 5

[0469] *delta-9 tetrahydrocannabinol

[0470] * cannabidiol

[0471] 10 *bPEI: branched polyethyleneimine; included in the “lipids” fraction for preparation purposes, according to the protocol of Example 1; although bPEI may not strictly be considered a lipid, but rather a transfection reagent to assist with endocytosis

[0472] Results were obtained from each group 11-15 using IVIS (as in Examples 3-5) to observe bioluminescence (relevant to luc mRNA) or fluorescence (relevant to GFP- 5 Cy5; the characteristic CY5 fluorescence Em peak is at 667 nm).

[0473] The results are shown in Figure 18 and confirm the trend already evidenced by the results of Example 5. The trend strongly suggests bypassing of the liver, i.e., mitigating or avoiding uptake of, and metabolism of, the delivered composition by the 10 liver. It further suggests preferential spleen and pancreas uptake, organs which may typically be difficult to target (especially when the liver sequesters API). Some bioluminescence is observed in the tail (site of injection).

[0474] Moreover, the following formulations showed a surprisingly high local 15 luciferase expression in target tissues (in this experiment, the pancreas and / or spleen) compared with the liver: 2C-LBSi-Tyr; 2C-LBSi-BPEI10-Tyr; 2C-LBSi-BPEI10-Tyr- SA; 2C-LBSi-THC. Again, the evidence points to “bypassing” the liver. Example 9 - Further murine model

[0475] In Example 9, the murine model of Example 4 was further developed.

[0476] An analogous protocol was followed to that of Example 4, except that different silicon-containing delivery platforms were used, and instead of siRNA, an investigative mRNA was used, namely exogenous firefly luciferase (Flue) or enhanced green fluorescent protein (eGFP) encoding mRNA. The silicon-containing delivery platforms were prepared and loaded with the appropriate mRNA and intravenously administered to wild-type mice. At 6 hours post-administration, the animals were sacrificed, and the organs harvested and snap-frozen for downstream characterisation of biodistribution.

[0477] The aim of the study was to understand the uptake and expression by different tissues of exogenous firefly luciferase or enhanced green fluorescent protein (eGFP) encoded mRNA, delivered silicon-containing delivery platforms, in murine tissue. The means used were reverse transcriptase quantitative PCR (RT-qPCR); complemented by fluorescence (ELISA, for eGFP) or luminescence (luciferase assay, for Flue) data.

[0478] The silicon-containing delivery platforms and mRNAs set out in Table 19 were used. See Examples 1 and 8 above for their formulations, which were prepared in the same way as the formulations detailed in Example 1.

[0479] The results are shown in Figures 19a-c and 20. They evidence bypassing of the liver, in that the splenic uptake of mRNA is considerably enhanced, from which it is inferred that liver sequestration of mRNA has been mitigated, allowing access to the spleen. Thus, targeting of a difficult-to-access organ is made possible.

[0480] ***

[0481] Where in the foregoing description, features or limitations are mentioned which have equivalents that are known, evident or foreseeable to those skilled in the art in the light of the present disclosure, then such equivalents are incorporated herein as if particularly set forth. Reference should be made primarily to the claims for determining the scope of the subject-matter of the present disclosure. The scope of protection sought by the present application further encompasses any such equivalents. It will also be appreciated by those skilled in the art that features or limitations of the disclosed subject-matter that are described as preferable, suitable, advantageous, convenient or the like may be optional and may not limit the scope of the independent claim(s) or the protection sought unless explicitly stated otherwise. Moreover, it is to be understood that such optional features or limitations, while of potential benefit in some implementations of the disclosed subject-matter, may be undesirable, and may therefore be absent or omitted in other implementations.

Claims

Claims1. A composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of treating cancer by miRNA administration to a human patient in need thereof.

2. A composition for use according to claim 1, wherein the composition further comprises (iii) the miRNA.

3. A composition for use according to claim 1 or claim 2, further comprising reducing or preventing the accumulation of the miRNA (which is an active pharmaceutical ingredient, “API”) in the liver.

4. A composition for use according to any preceding claim, further comprising promoting the accumulation of the miRNA (the API) in a target organ, especially the pancreas and / or spleen.

5. A composition for use according to any preceding claim, wherein the cancer is or comprises pancreatic cancer.

6. A composition for use according to claim 5, wherein the pancreatic cancer is or comprises pancreatic ductal adenocarcinoma.

7. A composition for use according to any preceding claim, wherein the cancer is or comprises cancer of the spleen.

8. A composition defined in accordance with any preceding claim, for use with one or more chemotherapeutic agents in a method of treating cancer in a human patient in need thereof.

9. A composition for use according to claim 8, wherein the composition is administered to the patient when the blood plasma level of the chemotherapeutic agents in the patient is below 0.1 pg / L.

10. A method of treating or preventing cancer, comprising administering to a human patient in need thereof a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) miRNA.

11. A method according to claim 10, further comprising reducing or preventing the accumulation of the miRNA (the API) in the liver.

12. A method according to claim 10 or claim 11, further comprising promoting the accumulation of the miRNA (the API) in a target organ, especially the pancreas and / or spleen.

13. A method according to any one of claims 10 to 12, wherein the cancer is or comprises pancreatic cancer.

14. A method according to claim 13, wherein the pancreatic cancer is or comprises pancreatic ductal adenocarcinoma.

15. A method according to any one of claims 10 to 14, wherein the cancer is or comprises cancer of the spleen.

16. A method according to any one of claims 10 to 15, further comprising administering one or more chemotherapeutic agents to the patient.

17. A method according to claim 16, wherein the composition is administered to the patient when the blood plasma level of the chemotherapeutic agents in the patient is below 0.1 pg / L.

18. A composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of reducing or preventing accumulation of an active pharmaceutical ingredient (API) in the liver of a human patient.

19. A composition for use according to claim 18, wherein the method further comprises promoting the accumulation of the API in a target organ; especially the pancreas and / or spleen.

20. A composition comprising (i) one or more lipids and (ii) particles comprising hydrolysable silicon, for use in a method of promoting accumulation of an active pharmaceutical ingredient (API) in the pancreas and / or spleen of a human patient in need thereof.

21. A composition for use according to claim 20, wherein the method further comprises reducing or preventing the accumulation of the API in the liver.

22. A composition for use according to any one of claims 19 to 21, wherein promoting the accumulation of the API in the pancreas and / or spleen is or comprises preventing or treating one or more diseases or disorders of the pancreas.

23. A composition for use according to claim 22, wherein the one or more pancreatic disorders is or comprise pancreatic cancer.

24. A composition for use according to claim 23, wherein the pancreatic cancer is or comprises pancreatic ductal adenocarcinoma.

25. A composition for use according to any one of claims 18 to 24, wherein the composition further comprises (iii) the API.

26. A composition for use according to any one of claims 19 to 25, wherein promoting the accumulation of the API in the pancreas and / or spleen is or comprises preventing or treating one or more diseases or disorders of the spleen.

27. A method of reducing or preventing accumulation of an active pharmaceutical ingredient (API) in the liver of a human patient in need thereof, comprising administering to the patient a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) the API.

28. A method according to claim 27, wherein the method further comprises promoting the accumulation of the API in a target organ, especially the pancreas and / or spleen.

29. A method of promoting the accumulation of an active pharmaceutical ingredient (API) in the pancreas and / or spleen of a human patient in need thereof, comprising administering to the patient a composition comprising (i) one or more lipids, (ii) particles comprising hydrolysable silicon and (iii) the API.

30. A method according to claim 29, wherein the method further comprises reducing or preventing the accumulation of the API in the liver.

31. A method according to any one of claims 28 to 30, wherein promoting the accumulation of the API in the pancreas and / or spleen is or comprises preventing or treating one or more diseases or disorders of the pancreas.

32. A method according to claim 31, wherein the one or more pancreatic disorders is or comprise pancreatic cancer.

33. A method according to claim 32, wherein the pancreatic cancer is or comprises pancreatic ductal adenocarcinoma.

34. A method according to any one of claims 28 to 33, wherein promoting the accumulation of the API in the pancreas and / or spleen is or comprises preventing or treating one or more diseases or disorders of the spleen.

35. A composition for use, or a method, according to any preceding claim, wherein (ii) the particles comprising hydrolysable silicon comprise at least 50 % w / w elemental silicon.

36. A composition for use, or a method, according to any preceding claim, wherein (ii) the particles comprising hydrolysable silicon are porous.

37. A composition for use, or a method, according to any preceding claim, wherein (ii) the particles comprising hydrolysable silicon have a mean diameter of about 10 to about 100 nm.

38. A composition for use, or a method, according to any preceding claim, wherein (i) the one or more lipids are or comprise one or more cationic lipids (e.g. DOTAP); one or more phospholipids (e.g. DOPE); and / or one or more polyethylene glycol (PEG) lipids (e.g. DSPE-PEG2000).

39. A composition for use, or a method, according to any preceding claim, wherein the composition further comprises (iv) one or more amino acids (especially glycine, arginine and / or tyrosine).

40. A composition for use, or a method, according to any preceding claim, wherein the composition further comprises (v) trehalose.

41. A composition for use, or a method, according to any preceding claim, wherein the method comprises reducing or preventing the accumulation of the API (which, when dependent on any one of claims 1 to 17, is the miRNA) in the liver, wherein reducing or preventing the accumulation of the API in the liver is or comprises preventing at least about 50 % of total API administered to the patient, from entering the liver.

42. A composition for use, or a method, according to any preceding claim, wherein the method comprises promoting accumulation of the API (which, when dependent on any one of claims 1 to 17, is the miRNA) in the pancreas, wherein promoting the accumulation of the API in the pancreas is or comprises effecting entry to the pancreas of at least about 50 % of total API administered to the patient.

43. A composition for use, or a method, according to any preceding claim, wherein the method comprises promoting accumulation of the API (which, when dependent on any one of claims 1 to 17, is the miRNA) in the spleen, wherein promoting the accumulation of the API in the spleen is or comprises effecting entry to the spleen of at least about 50 % of total API administered to the patient.

44. A composition for use, or a method, according to any preceding claim, wherein the method comprises promoting accumulation of the API (which, when dependent on any one of claims 1 to 17, is the miRNA) in the pancreas and spleen, wherein promoting the accumulation of the API in the pancreas and spleen is or comprises effecting the distribution of an amount of at least 50 % of total API administered to the patient, to a group of organs comprising or consisting of the pancreas and the spleen.

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