WO2025019817A1 - Multi-specific self-assembled drug-free macromolecular t-cell engagers - Google Patents

Abstract

Disclosed herein is a first conjugate comprising a first targeting moiety that is adapted to specifically bind to a first antigen on the surface of a target effector cell and a first morpholino oligonucleotide; and a plurality of additional conjugates comprising a targeting moiety that is adapted to bind to a second antigen that is on the surface of a target B-cell and a morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. Also described herein are kits comprising the conjugates and methods for using the conjugates and kits.

Description

MULTI-SPECIFIC SELF-ASSEMBLED DRUG-FREE MACROMOLECULAR T-CELL ENGAGERS

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No. 63/514,666, filed July 20, 2023, U.S. Provisional Patent Application No. 63/593,469, filed October 26, 2023, U.S. Provisional Patent Application No. 63/631 ,095, filed April 8, 2024, U.S. Provisional Patent Application No. 63/663,907, filed June 25, 2024, each of which is incorporated herein by reference in its entirety.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

[0002] This invention was made with government support under grants R01 GM095606 and R01 CA246716 awarded by the National Institutes of Health. The government has certain rights in the invention.

REFERENCE TO SEQUENCE LISTING

[0003] This application was filed with a Sequence Listing XML in ST.26 XML format accordance with 37 C.F.R. § 1.831 and PCT Rule 13ter. The Sequence Listing XML file submitted in the USPTO Patent Center, “026389-0022-W001 ,” was created on July 19, 2024, contains 54 sequences, has a file size of 47.7 Kbytes, and is incorporated by reference in its entirety into the specification.

FIELD

[0004] This disclosure relates to a first conjugate comprising a first targeting moiety that is adapted to specifically bind to a first antigen on the surface of a target effector cell and a first morpholino oligonucleotide; and a plurality of additional conjugates comprising a targeting moiety that is adapted to bind to a second antigen that is on the surface of a target B-cell and a morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. Also disclosed herein are kits comprising the conjugates and methods for using the conjugates and kits.

INTRODUCTION [0005] Bispecific, T cell-engaging antibody constructs allow for T-cell recruitment and activation against antigen-expressing cancer cells. Current T-cell recruiting approaches bridge the synapse between a target cancer cell and an effector T-cell using constructs capable of binding antigens on both cells simultaneously. At the cell-to-cell interface, clustering of bispecific molecules initiates T-cell activation, polarization, and degranulation of cytolytic granules towards the target cancer cell. The first FDA approved bispecific T-cell engager was blinatumomab - a fusion protein comprised of an a-CD19 short chain variable fragment (scFv) and an a-CD3 scFv fused together and expressed as a single molecule. Blinatumomab can engage with target malignant B-cells through CD19 proteins on the cancer cell surface, and simultaneously engage with cytotoxic T-cells through CD3 proteins in the T-Cell Receptor (TCR) complex. Blinatumomab induces a TCR-like activation response in effector T-cells and directs their cytotoxicity towards CD19(+) cancer cells. More recently, numerous bispecific T-cell engagers have been FDA approved for the treatment of multiple myeloma and lymphoma, highlighting that T-cell directing therapies are a highly effective therapeutic approach.

[0006] The efficacy of T-cell recruiting strategies is contingent upon expression of the target antigen on the surface of the cancer cell. Remission duration is dependent on the therapy’s ability to recruit cytotoxic T cells to antigen-d/7n phenotypes of the heterogenous cancer cell population. Serendipitous downregulation of target antigen molecules on the cancer cell surface hampers the effectiveness of T-cell therapies and allows the cancer cells to avoid destruction. Incomplete removal of antigen-d/m cell subsets leads to only a partial response and relapse of a malignant population with decreased or absent antigen expression leading to resistance to the therapy. Dual- and multi-targeted T-cell recruiting strategies look to combat problems of relapse by broadening cancer cell targeting. The simultaneous targeting of two or more antigens on the cancer cell surface increases the selective therapeutic pressure and decreases occurrence of antigen-negative relapse. Immunotherapeutic platforms capable of multi-targeted T-cell recruitment are under development in preclinical and clinical trials. Examples include multi-specific CAR-T, immunoliposomes, antibody constructs, and DNA- origami T-cell engagers.

[0007] Thus, there is a need for a T-cell recruiting methodology designed for simple customizability to treat cancer cells based on their unique antigen expression profiles.

SUMMARY [0008] In an aspect, the disclosure relates to a kit comprising: (i) a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; (ii) a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of a target B-cell; and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell. In an embodiment, the plurality of additional conjugates further includes at least a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the fourth targeting moiety is adapted to bind to a fourth antigen that is on the surface of the target B-cell. In another embodiment, the second morpholino oligonucleotide and the third morpholino oligonucleotide have the same nucleotide sequence. In another embodiment, the second and third morpholino oligonucleotides are both 95% complementary to the first morpholino oligonucleotide. In another embodiment, the second and third morpholino oligonucleotides are both 100% complementary to the first morpholino oligonucleotide. In another embodiment, the first targeting moiety is coupled to the first morpholino oligonucleotide by a first linker, the second targeting moiety is coupled to the second morpholino by a second linker and the third targeting moiety is coupled to the third morpholino oligonucleotide by a third linker. In another embodiment, the first linker, the second linker, and the third linker are each independently from about 10 A to about 100 A in length. In another embodiment, one or more of the first linker, the second linker, and the third linker comprises polyethylene glycol (PEG) or succinimidyl-4-(N- maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), and wherein each of the first linker, the second linker, and the third linker have either the same or a different molecular weight relative to one another. In another embodiment, the first, second and third morpholino oligonucleotides are each independently from about 10 nucleotides to about 30 nucleotides in length. In another embodiment, the first targeting moiety is a first antibody or a first Fab’ fragment of the first antibody, and wherein the first antigen is selected from the group consisting of CD3, CD16, CD14, and CD64. In another embodiment, the first antibody is selected from the group consisting of anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), antiCDF (clone: UCHM-1), and anti-CD64 (clone 10.1). In another embodiment, the first antigen is CD3. In another embodiment, the first antibody is anti-CD3 (clone: UCHT-1). In another embodiment, the second targeting moiety is a second antibody or a second Fab’ fragment of the second antibody and the third targeting moiety is a third antibody or a third Fab’ fragment of the third antibody, and wherein the second and third antigens are each independently selected from the group consisting of CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38, CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1, BCMA, SLAMF7 (CS-1), and GPRC5D. In another embodiment, the second and third antibodies are each independently selected from the group consisting of UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, and Talquetamab. In another embodiment, the second and third antigens are each independently selected from the group consisting of CD3, CD19, CD20, CD38, CD52, BCMA, and SLAMF7 (CS-1). In another embodiment, the second and third antibodies are each independently selected from the group consisting of UCHT-1 , SJ25-C1 , Rituximab, Obinutuzumab, Daratumumab, Isatuximab, Alemtuzumab, Belantamab, and Elotuzumab. In another embodiment, the second and third conjugates are co-formulated. In another embodiment, the second and third conjugates are further co-formulated with the first conjugate. In another embodiment, the plurality of additional conjugates further includes at least a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide; wherein the fourth targeting moiety is adapted to bind to a fourth antigen that is on the surface of the target B-cell; wherein the fourth conjugate is co-formulated with the second and third conjugates. In another embodiment, the nucleotide sequence of the first morpholino oligonucleotide is SEQ ID NO: 2 and the nucleotide sequence of the second and third morpholino oligonucleotides is SEQ ID NO: 1. In another embodiment, the nucleotide sequence of the fourth morpholino oligonucleotide is SEQ ID NO: 1. [0009] In a further aspect, the disclosure relates to a method of inducing apoptosis of a target B-cell, the method comprising: (i) contacting a population of cells with a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; and (ii) contacting the population of cells with a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of the target B-cell; a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell; and wherein the first morpholino oligonucleotide hybridizes with the second morpholino oligonucleotide, and the target effector cell induces apoptosis of the target B-cell. In an embodiment, the population of cells are in a subject. In another embodiment, the subject is a mammalian subject having cancer or an autoimmune disorder. In another embodiment, the second morpholino oligonucleotide and the third morpholino oligonucleotide have the same nucleotide sequence. In another embodiment, the first targeting moiety is coupled to the first morpholino oligonucleotide by a first linker, the second targeting moiety is coupled to the second morpholino by a second linker and the third targeting moiety is coupled to the third morpholino oligonucleotide by a third linker. In another embodiment, the first linker, the second linker and the third linker are each independently is from about 10 A to about 100 A in length. In another embodiment, one or more of the first linker, the second linker and the third linker comprises polyethylene glycol (PEG) or succinimidyl-4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), and wherein each of the first linker, the second linker and the third linker have either the same or a different molecular weight relative to one another. In another embodiment, the first, second and third morpholino oligonucleotides are each independently from about 10 nucleotides to about 30 nucleotides in length. In another embodiment, the first targeting moiety is a first antibody or a first Fab’ fragment of the first antibody, and wherein the first antigen is selected from the group consisting of CD3, CD16, CD14, and CD64. In another embodiment, the first antibody is selected from the group consisting of anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), antiCDF (clone: UCHM-1), and anti-CD64 (clone 10.1). In another embodiment, the first antigen is CD3. In another embodiment, the first antibody is anti-CD3 (clone: UCHT-1). In another embodiment, the second targeting moiety is a second antibody or a second Fab’ fragment of the second antibody and the third targeting moiety is a third antibody or a third Fab’ fragment of the third antibody, and wherein the second and third antigens are each independently selected from the group consisting of CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38. CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1, BCMA, SLAMF7 (CS-1), and GPRC5D. In another embodiment, the second and third antibodies are each independently selected from the group consisting of UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, and Talquetamab. . In another embodiment, the second and third antigens are each independently selected from the group consisting of CD3, CD19, CD20, CD38, CD52, BCMA and SLAMF7 (CS-1). In another embodiment, the second and third antibodies are each independently selected from the group consisting of UCHT-1 , SJ25-C1 , Rituximab, Obinutuzumab, Daratumumab, Isatuximab, Alemtuzumab, Belantamab and Elotuzumab. In another embodiment, the nucleotide sequence of the first morpholino oligonucleotide is SEQ ID NO: 2 and the nucleotide sequence of the second and third morpholino oligonucleotides is SEQ ID NO: 1. In another embodiment, the first conjugate and the plurality of additional conjugates are administered to the subject in an amount ranging from about 1 pg/kg to about 500 mg/kg. In another embodiment, the ratio of the amount of first conjugate and the amount of the plurality of additional conjugates administered to the subject ranges from about 1 ,000:1 to about 1 :1 ,000. In another embodiment, the second and third conjugates are co-administered to the subject before the first conjugate is administered to the subject, and wherein the first conjugate is administered to the subject from about 0.5 hours to about 72 hours after the second and third conjugates were administered to the subject.

[00010] The disclosure provides for other aspects and embodiments that will be apparent in light of the following detailed description and accompanying figures.

BRIEF DESCRIPTION OF THE DRAWINGS [00011] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.

[00012] FIG. 1 is a schematic representation of effector cell recruitment and activation against malignant or autoactivated B-cells.

[00013] FIG. 2 is a schematic representation of an exemplary two-component effector cell activator therapy against malignant or autoreactive B-cells.

[00014] FIGS. 3A, 3B, 3C, and 3D illustrate the synthetic route for making antibody binding fragment (Fab’)-morpholino oligonucleotide conjugates (Fab’-MORF1/2). The whole antibody is enzymatically cleaved below the hinge region cysteine residues using pepsin. Resulting F(ab’)2 intermediates are reduced using TCEP to generate desired thiol-active Fab’-SH species. In parallel to the reduction reaction, 3’-amine functionalized MORF strands are conjugated to the bi-functional SM(PEG)2 linker by N-hydroxysuccinimide ester-amine coupling. The resulting maleimide-functionalized MORF-PEG2-maleimide species is conjugated to the thiol-activated Fab’-SH species by thiol-maleimide click chemistry. Final products are purified using ultracentrifugation and characterized for purity (size exclusion chromatography), mass (mass spectroscopy), MORF-to-Fab’ substitution ratio (bicinchoninic acid and UV-Vis), and dimerization efficiency (UV-Vis, size exclusion chromatography, and dynamic light scattering). FIG. 3A is a schematic of the synthetic route for making antibody binding fragment (Fab’)- morpholino oligonucleotide conjugates. FIG. 3B is a graph of size exclusion chromatograms of the enzymatic digestion of whole antibody into the F(ab’)₂ intermediate species over time performed with a Superdex 200 protein column in PBS with a flow rate of 0.4 mL min'¹. FIG. 3C is a graph of size exclusion chromatograms of whole antibody, F(ab’)₂, and Fab’ intermediate products performed with a Superdex 200 protein column in PBS with a flow rate of 0.4 mL min^-1. FIG. 3D is a graph of size exclusion chromatograms of CD20-directed, CD38-directed, SLAMF7(CS-1)-directed, and BCMA-directed B-cell binding motifs performed with a Superdex 200 protein column in PBS with a flow rate of 0.4 mL min'¹.

[00015] FIG. 4 is a graph of size exclusion chromatogram of T-cell engager, Fab’_CD3-MORF2 along with the mass spectrometry pictograph performed with a Superdex 200 protein column in PBS with a flow rate of 0.4 mL min'¹. Size exclusion chromatography of the final, purified T-cell engager product, Fab’cD3-MORF2 with the accompanying time of flight positive electron spray mass spectrometry (TOF MS ES+). The chromatogram suggests a 95% pure product supported by the mass spectrometry plot showing a single MORF2 substituted Fab’ product.

[00016] FIGS. 5A, 5B, and 5C show further spontaneous self-assembly of the T-cell engager, Fab’_CD3-MORF2, with Fab’_CD2o-MORF1. FIG. 5A is a graph depicting size exclusion of premixture of T-cell engager and CD20-directed B-cell engager. FIG. 5B is a graph depicting the hypochromic effect of MORF1/MORF2 hybridization on UV-Vis measuring absorbance at 260 nm. FIG. 5C is a graph showing dynamic light scattering of Fab’_CD3-MORF2, Fab’_CD2o- MORF1, and the premixed solution.

[00017] FIG. 6 is a graph of a size exclusion chromatogram of CD19-directed B-cell binding motifs performed with a Superdex 200 protein column in PBS with a flow rate of 0.4 mL min'¹.

[00018] FIG. 7 is an image of light microscopy (top) and confocal microscopy (bottom) of cocultured Raji B-cell and Jurkat T-cells. Cells were treated with Cy5-labeled Fab’cD3-MORF2 (red) and Cy3-labeled Fab’_CD2o-MORF1 (green) for 1 h. Overlap (yellow), designated by white arrows, shows co-localization of fluorophores at the cell-to-cell synapse.

[00019] FIGS. 8A, 8B, and 8C show expression profiling and cell-specific T-cell activation of leukemia (HL-60), lymphoma (Raji), and myeloma (MM.1S) cell lines using either CD38-, CD20- , BCMA-, or SLAMF7-directed MATCH. Apoptosis of target B-cells was quantified after 2 h coculture of T-cells and target cells in a 1 :1 ratio using APC-annexin V and propidium iodide and quantified using flow cytometry. Cancer Cell Line Antigen Profiling. Immortalized cell lines including: i) B-cell lines of Raji (NHL), Daudi (NHL), MM.1S (MM), RPMI 8226 (MM), KMS-12- BM (MM), ANBL-6 (MM), U266 (MM); ii) the T cell line Jurkat (ALL); and ill) the myeloid line HL- 60 (AML) were examined for their CD20, CD38, BCMA, and SLAMF7 expression levels. Cells were treated with fluorescently labeled primary antibodies and analyzed using cell sorting.

Antigen expression was quantitated by normalizing geometric mean averages of labeled cells to untreated control. FIG. 8A is a graph of expression profiling of leukemia (HL-60). FIG. 8B is a graph of expression profiling of lymphoma (Raji). FIG. 8C is a graph of expression profiling of myeloma (MM.1S).

[00020] FIGS. 9A, 9B, 9C, 9D, 9E, 9F, 9G, and 9H depict the same cohort of T-cells (2 x 10⁶) Activating a Cohort of Naive T-Cells Against Multiple Cancers Using Cell-Specific MATCH. The same batch of naive, healthy donor T-cells were sequentially rechallenged with cancer cells over time. For the first challenge, T cells (2 x 10⁶) were added to a 6-well plate along with Raji B-cells (2 x 10⁵) in 4 mL RPMI 1640 cell culture medium. Cells were incubated with or without a premixed dose of Fab’cD2o-MORF1/Fab’_CD3-MORF2 (50 nM) for 24 h at 37 °C. After 24 h, an aliquot (1 mL) was removed and stained using APC-CD19 antibody. Number of CD19(+) cells in the untreated co-culture were qualitatively compared to the MATCH -treated cells. Remaining co-cultures were collected, washed with PBS, and resuspended in fresh RPMI 1640 culture medium. The second challenge commenced by adding MM.1S (1.5 x 10⁵) cells to each well and bringing the total volume to 4 mL. Cells were incubated with or without a premixed dose of Fab’BCMA- MORF1/Fab’cD3-MORF2 (50 nM) for 24 h at 37 °C. After 24 h, aliquot (1 mL) was removed and stained using PE-CD10 antibody. Number of CD10(+) cells in the untreated coculture were compared to the MATCH-treated cells. Remaining co-cultures were collected, washed with PBS, and resuspended in fresh RPMI 1640 medium. The third challenge commenced with the addition of HL-60 (1 .12 x 10⁵) cells to each well. Cells were incubated with or without a premixed dose of Fab’cD38-MORF1/Fab’cD3-MORF2 (50 nM) for 24 h at 37 °C. After 24 h, aliquots were stained with PE-CD33 antibody to compare number of HL-60 in untreated co-cultures to the MATCH-treated cells. The entire experiment was repeated using blinatumomab as the therapeutic agent dosed at 50 nM. All experiments were replicated two times. FIG. 9A is a schematic of the untreated control growth. FIG. 9B is a graph of CD19 expression for lymphoma in an untreated co-culture. FIG. 9C is a graph of CD10 expression for myeloma in an untreated co-culture. FIG. 9D is a graph of CD33 expression for leukemia in an untreated co-culture. FIG. 9E is a schematic of the MATCH control growth. FIG. 9F is a graph of CD19 expression for lymphoma in a CD20 MATCH culture. FIG. 9G is a graph of CD10 expression for myeloma in a BCMA MATCH culture. FIG. 9H is a graph of CD33 expression for leukemia in a CD38 MATCH culture.

[00021] FIG. 10 is an image of confocal microscopy of Raji B-cells co-cultured with healthy donor T-cells with or without CD20-directed MATCH therapy for 1 h. White boxes indicate areas of interest.

[00022] FIG. 11 is an image of confocal microscopy of T-cells (blue) inducing perforin pores (green) in target B-cell membranes (red). To detect the presence of perforin at cell interfaces in response to either MATCH or blinatumomab treatment, 2.0 x 10⁵ healthy human T cells and 2.0 x 10⁵ Raji cells were seeded in a 24-well plate in 400 pL RPMI 1640 medium and incubated for 4 h under standard incubation conditions. Cells were then washed and immunostained with aPerforin-FITC, aCD4-PE/aCD8-PE, and either aCD20-APC (for untreated and blinatumomab- treated cells) or aCD19-APC (for aCD20 MATCH-treated cells). After washing, 1 .0 x 10⁶ cells per sample were loaded into the chamber and examined by confocal microscopy.

[00023] FIGS. 12A and 12B show calcium influx via perforin pore formation in target cell membranes. CD20-directed MATCH (left) and blinatumomab control (right). To quantify calcium influx in target cells in response to either MATCH or blinatumomab treatment, 2.0 x 10⁵ healthy human T cells and 2.0 x 10⁵ Raji cells were seeded in a 24-well plate in 400 pL RPMI 1640 medium supplemented with (concentration of calcium rich) and incubated for 2 h under standard incubation conditions. Cells were then immunostained with aCD4-PE/aCD8-PE, and either aCD20-APC (for untreated and blinatumomab-treated cells) or aCD19-APC (for aCD20 MATCH-treated cells). After washing, Fluo-3 AM fluorescence in target cells of each group was quantified using flow cytometry. FIG. 12A is a graph of Flu-3 AM versus counts of target cells for CD20 MATCH culture. FIG. 12B is a graph of Flu-3 AM versus counts of target cells for Blinatumomab culture as a culture.

[00024] FIGS. 13A, 13B, 13C, 13D, 13E, 13F, and 13G are graphs showing MATCH- Induced mitochondrial depolarization in target cancer cells. Mitochondrial membrane integrity was tested using the mitochondrial membrane potential sensor JC-1 (Thermo Scientific). T cell- to-target cell ratio of 1 :1 in a 48-well plate in 400 pL RPMI 1640 media were dosed with or without CD20-directed MATCH (50 nM) for 4 h at 37 °C. After 4 h, wells were collected, washed, and stained with JC-1 (4 pM) for 30 min at 37 °C. For positive depolarized control, untreated cells were treated with CCCP (0.5 pM) and incubated simultaneously with JC-1 for 30 min. After all JC-1 treatments, cells were washed analyzed using flow cytometry. FIG. 13A is a graph of flow cytometry data for an untreated co-culture. FIG. 13B is a graph of flow cytometry data for a CCCP culture. FIG. 13C is a graph of flow cytometry data for a Fab’_CD2o-MORF1 culture. FIG. 13D is a graph of flow cytometry data for a CD20 MATCH culture. FIG. 13E is a graph of flow cytometry data for a Blinatumomab culture. FIG. 13F is a graph of flow cytometry data for a Fab’cD3-MORF2 culture.

[00025] FIG. 14 is an image of confocal microscopy of co-cultured healthy, naive T-cells with Raji B-cells investigating CD20-directed MATCH induced mitochondrial depolarization in target B-cells. The mitochondrial membrane molecule JC-1 was used to detect depolarized membranes. Healthy membranes (red), depolarized membranes (green) and overlay (yellow) of CD20-directed MATCH treated cells (top panels) are compared to blinatumomab treated cells (bottom panels). [00026] FIGS. 15A and 15B are graphs representing caspase activation within target Raji B- cells when treated with CD20-directed MATCH compared to blinatumomab. To directly quantify caspase 3 activation in response to MATCH or blinatumomab treatment, 2.0 x 10⁵ healthy human T cells and 2.0 x 10⁵ Raji cells were seeded in a 24-well plate in 400 pL RPMI 1640 medium. The caspase 3 substrate PhiPhiLux®GiD₂ was administered to each sample and the cells incubated for 4 h under standard incubation conditions. Cells were then immunostained with aCD4-PE/aCD8-PE, and either aCD20-APC (for untreated and blinatumomab-treated cells) or aCD19-APC (for aCD20 MATCH-treated cells). After washing, substrate cleavage (as measured by emission on the FITC channel) was quantified for each group using flow cytometry. FIG. 15A is a graph of caspase activation for MATCH-induced cultures. FIG. 15B is a graph of flow cytometry data collected after administering caspase 3 substrate PhiPhiLux®GiD2.

[00027] FIG. 16 is a graph of CD20-directed MATCH in combination with either a pan protease inhibitor or an anti-FasL antibody to determine contributions of caspases and Fas- FasL interactions to overall MATCH-induced apoptosis. Compared to blinatumomab. To quantify the contribution of Fas-FasL interaction in target cell clearance during either MATCH or blinatumomab treatment, 2.0 x 10⁵ health human T cells and 2.0 x 10⁵ Raji cells were seeded in a 24-well plate in 400 pL RPMI 1640 medium. For aCD20 MATCH and blinatumomab, an additional treatment group was simultaneously treated with aFasL antibody blockade and all cells incubated for 4 h under standard incubation conditions. Cells were then immunostained with aAnnexin-FITC, DAPI, and either aCD20-APC (for untreated and blinatumomab-treated cells) or aCD19-APC (for aCD20 MATCH-treated cells). After washing, the fraction of apoptotic target cells was quantified by flow cytometry. In particular, the difference in apoptotic cell fraction between treatment groups with and without aFasL antibody blockade was quantified. To inhibit granzyme B activity during MATCH or blinatumomab treatment, excess human T cells were treated with (concentration and name of cocktail and time/conditions of incubation). After washing, 2.0 x 10⁵ of these pre-treated healthy human T cells and 2.0 x 10⁵ Raji cells were seeded in a 24-well plate in 400 pL RPMI 1640 medium. As a control, each treatment group was replicated using T cells which had not been pre-treated protease inhibitor cocktail. All cells incubated for 4 h under standard incubation conditions and were then washed and immunostained with aAnnexin-FITC, DAPI, and either aCD20-APC (for untreated and blinatumomab-treated cells) or aCD19-APC (for aCD20 MATCH-treated cells). After washing, percentage of apoptotic target cells was determined by flow cytometry. In particular, the difference in apoptotic cell fraction between treatment groups with and without the protease inhibitor cocktail was quantified.

[00028] FIGS. 17A, 17B, 17C, 17D, 17E, 17F, 17G, 17H, and 171 depict flow cytometry data of CD20-directed MATCH (compared to blinatumomab) in combination with either a pan protease inhibitor or an anti-FasL antibody to determine contributions of caspases and Fas- FasL interactions to overall MATCH-induced apoptosis. FIG. 17A is a graph of flow cytometry for an untreated culture. FIG. 17B is a graph of flow cytometry for Fab’_CD2o-MORF1. FIG. 17C is a graph of flow cytometry for Fab’cD3-MORF2. FIG. 17D is a graph of flow cytometry for CD20. FIG. 17E is a graph of flow cytometry for MATCH + Protease Inhibitors. FIG. 17F is a graph of flow cytometry for MATCH+FasL Blockade. FIG. 17G is a graph of flow cytometry for Blinatumomab. FIG. 17H is a graph of flow cytometry for Blinatumomab + Protease Inhibitors. FIG. 171 is a graph of flow cytometry for Blinatumomab + FasL Blockade.

[00029] FIGS. 18A, 18B, and 18C are graphs of mitochondrial depolarization (JC-1 aggregation), caspase inhibition (Z-VAD-FMK), and reactive oxygen species (H2DCFDA) generated from CD20-directed MATCH on Raji B-cells after 4 h. Levels of reactive oxygen species in the target cell cytosol after 4 h MATCH treatments were quantified by oxidation of 2’,7’-dichlorodihydrofluorescein diacetate (H₂DCFDA). Target cells were treated co-cultured in a T cell-to-target cell ratio of 1 :1 in a 48-well plate in 400 L RPMI 1640 media were dosed with or without CD20-directed MATCH (50 nM) for 4 h at 37 °C. After treatment, cells were incubated with H2DCFDA (5 |JM) for 30 min at 37 °C. Cells were washed with PBS and analyzed using flow cytometry. FIG. 18A is a graph of mitochondrial depolarization in various cultures. FIG. 18B is a graph of caspase activation in various cultures. FIG. 18C is a graph of reactive oxygen species among an untreated co-culture and CD20 MATCH culture.

[00030] FIGS. 19A, 19B, and 19C are graphs of cell viability assays on MM.1S cells using monospecific MATCH therapies versus a trispecific combinatorial MATCH therapy incorporating CD38, BCMA, and SLAMF-7. Curves were obtained using CCK-8 assay in 96-well plates. Cocultures of T-cells and Raji cells were exposed to a series of MATCH concentrations for 24 h. Dose Response of Multi-Targeted MATCH. The dose response of Multi-specific MATCH was assayed using a metabolic viability approach following a Chou-Talalay combination therapy setup. Proper Chou-Talalay method requires prior knowledge of EC50 values of individual therapies to construct a cocktail of combinational therapies at fold increases or fold decreases of the respective EC50 values for each therapy in the cocktail. MM.1S cells were triple positive, thus, three single target dose response curves were attained for BCMA-, SLAMF7-, and CD38- directed MATCH, respectively. Brielfy, a co-culture of naive, healthy donor T-cells with target myeloma cells (MM.1S) were plated in a 96-well plate in a 1:1 ratio (1x10⁴ of each cell per well). Cell co-cultures were dosed with serial dilutions of either BMCA-, SLAMF7-, or CD38-directed MATCH, from a dose range of 10 pM to 100 nM, for 24 h at 37 °C. After 24 h, CCK-8 (10 piper well) was added and incubated for 2 h at 37 °C. Absorbance at 460 nm was measured and plotted using GraphPad Prism using non-linear least squares fit. ECso values for each single target therapy were attained and further used in the Chou-Talalay combination evaluation method. Briefly, serial dilutions above and below the respective ECso values of each B-cell engaging Fab’-MORF1 were combined and dosed such that the final Fab’-MORF1 cocktail MORF1 concentration was combined with a one-to-one ratio of the complementary T-cell engager Fab’cD3-MORF2 such that [MORF1]=[MORF2], For the multi-specific MATCH experiment, naive healthy T-cells were co-cultured in a 1 -to-1 ratio with MM.1S cells (1x10⁴ of each cell per well) in a 96-well plate. Premixed cocktails of Fab’_BcviA-MORF1 , Fab’_SLAMF7- MORF1, Fab’cD3s-MORF1, and Fab’_CD3-MORF2 were titrated and added to each well. The cells were incubated for 24 h at 37 °C. After 24 h, CCK-8 (10 pL per well) was added and incubated for 2 h at 37 °C. Absorbance at 460 nm was measured and plotted using GraphPad Prism using non-linear least squares fit. FIG. 19A is a graph of the effect Fab’scMA-MORFI concentration on BCMA T-Cell activation. FIG. 19B is a graph of the effect Fab’cs-i-MORF1 concentration on CS-1 T-Cell activation. FIG. 19C is a graph of the effect Fab’_CD3s-MORF1 concentration on CD38 T-Cell activation.

[00031] FIGS. 20A, 20B, 20C, 20D, and 20E are graphs of consecutive MATCH dosing versus premixed administration. Co-culture of T-cells and Raji B-cells in a 3:1 B cell-to-T cell ratio. In consecutive dosing, Raji cells were treated for 1 h with Fab’_CD2o-MORF1 , washed, resuspended in fresh media then co-cultured with T-cells where finally the T-cell engager was titrated in at denoted concentrations. Blinatumomab was used as control. B-cell depletion after 24 h was measured using flow cytometry (left axes) and T-cell exhaustion, measured as PD-1 expression increase, was measured using flow cytometry (right axes). FIGS. 20A is a graph of B-cell depletion for CD20 MATCH consecutive administration. FIG. 20B is a graph of B-cell depletion for premixed conjugates. FIG. 20C is a graph of B-cell depletion for CD19 MATCH consecutive administration. FIG. 20D is a graph of B-cell depletion for CD19 MATCH premixed conjugates. FIG. 20E is a graph of the B-cell depletion of Blinatumomab, used as a control. [00032] FIGS. 21A and 21B show a 2-dimensional dose response of healthy, naive T-cells and Raji (luciferase) B-cells co-cultured in 96-well plates for 24 h. Bioluminescence measured by luciferin administration after 5 min using IVIS imager. FIG. 21 A shows the dose response for a premixture 1 -to-1 E:T ratio. FIG. 21B shows the dose response for a consecutive 1-to-1 E:T ratio.

[00033] FIGS. 22A, 22B, 22C, 22D, 22E, 22F, 22G, 22H, 22I, 22J, 22K, 22L, and 22M are graphs of T-cell cytokines released after 24 h treatment of CD19- or CD20-directed MATCH compared to blinatumomab at doses of 50 nM, 10 nM, 5 nM, 1 nM, and 500 pM. Quantified using the T-cell/NK-cell LegendPLEX kit from BioLegend. LEGENDplex™ Human CD8/NK Panel V02 kits (BioLegends) were used to determine levels of cytokine release during MATCH treatment, according to manufacturer’s protocol. Five treatment regimens were assay (blinatumomab, aCD19 MATCH simultaneous administration, aCD19 MATCH consecutive administration, aCD20 MATCH simultaneous administration, and aCD20 MATCH consecutive administration). Each treatment was assayed at 5 treatment doses (50 nM, 10 nM, 5 nM, 1nM, and 500 pM, with equal doses of both Fab’-MORF conjugates used for all MATCH treatments). To prepare samples for cytokine release analysis, 1.0 x 10⁵ T-cells and 1.0 x 10⁵ target cells were seed into a 48-well plate. For consecutively-administered MATCH treatment groups, B cells were treated with Fab’_CDi9-MORF1 or Fab’_RTx-MORF1 for 30 minutes and then seeded in the plate. For simultaneously-administered MATCH treatment groups and blinatumomab, B cells were seeded in the plate with T cells without pre-treatment (and then the appropriate dose of Fab’cDi9-MORF1 or Fab’_RTx-MORF1 was administered for the MATCH groups).

Blinatumomab or Fab’_CD3-MORF2 was then administered as appropriate, and the cells incubated for 24 h. All samples were then centrifugated and the supernatant was collected and prepared for analysis. The remaining cells were washed and immunostained with aCD20 (for aCD19 MATCH- and blinatumomab-treated groups) or aCD19 (for aCD20 MATCH-treated groups), and cell counts were quantified by flow cytometry. FIG. 22A is a graph of the flow cytometry for IL-2. FIG. 22B is a graph of the flow cytometry for IL-4. FIG. 22C is a graph of the flow cytometry for IL-6. FIG. 22D is a graph of the flow cytometry for IL- 10. FIG. 22E is a graph of the flow cytometry for IL-17A. FIG. 22F is a graph of the flow cytometry for IFN-y. FIG. 22G is a graph of the flow cytometry for TNF-a. FIG. 22H is a graph of the flow cytometry for sFasL. FIG. 22I is a graph of the flow cytometry for granulysin. FIG. 22J is a graph of the flow cytometry for perforin. FIG. 22K is a graph of the flow cytometry for granzyme A. FIG. 22L is a graph of the flow cytometry for granzyme B. FIG. 22M is a graph of the flow cytometry for sFas. [00034] FIGS. 23A, 23B, 23C, 23D, 23E, and 22F show T-cell cytokine release and B-cell ablation efficacy of 2:1 B-cell to T-cell ratio with CD19- and CD20-directed MATCH, dosed as both a premixture and consecutively, compared to blinatumomab. FIG. 23A shows depletion of target cells after a 24 hour treatment at varying doses of CD19 MATCH, CD20 MATCH, and Blinatumomab. FIG. 23B is a graph of CD20 MATCH premix ablation after 24 at the boxed dose (5 nM) of FIG. 23A. FIG. 23C is a graph of CD20 MATCH consecutive ablation after 24 at the boxed dose (5 nM) of FIG. 23A. FIG. 23D is a graph of CD19 MATCH premix ablation after 24 at the boxed dose (5 nM) of FIG. 23A. FIG. 23E is a graph of CD19 MATCH consecutive ablation after 24 at the boxed dose (5 nM) of FIG. 23A. FIG. 23F is a graph of Blinatumomab ablation after 24 at the boxed dose (5 nM) of FIG. 23A.

[00035] FIGS. 24A, 24B, 24C, and 24D show T-cell cytokine release and B-cell ablation efficacy of 2:1 B-cell to T-cell ratio. CD19- and CD20-directed MATCH, dosed as both a premixture and consecutively, compared to blinatumomab at the 5 nM dose. FIG. 24A is a graph of T-cell cytokine release for interleukin levels. FIG. 24B is a graph of T-cell cytokine release for cytolytic molecule levels. FIG. 24C is a graph of T-cell cytokine release for IFN-y. FIG. 24D is a graph of T-cell cytokine release for TNF-a.

[00036] FIGS. 25A and 25B are graphs of In vivo validation of CD20-directed MATCH on xenograft non-Hodgkin’s lymphoma in SCID mice. Inoculation and dose schedule. Single dose MATCH (0.6 pg; blue) was compared head-to-head against single dose blinatumomab (0.6 pg; green). Additionally, a cohort of mice were treated with single-dose MATCH (60 pg; orange). Mouse survival curve (n=3). Untreated control mice (Raji alone; red, and Raji + T Cell xenograft with no treatment; brown) presented with hind-limb paralysis by week three. MATCH at 1 nanomole dose (orange) saw 3/3 mice live >75 days with no evidence of disease. Equivalent doses of MATCH to blinatumomab (0.6 pg) observed trending improvement in survival in the MATCH cohort over blinatumomab, although not significant (p=0.08). FIG. 25A is a schematic of the dose schedule for the MATCH dosing of C.B-17 mice. FIG. 25B is a graph showing mouse survival over time after MATCH dosing of untreated and Blinatumomab treated mice.

[00037] FIGS. 26A, 26B, 26C, and 26D are graphs of ex vivo bone marrow analysis of CD10(+)/CD19(+) cells of MATCH (0.6 pg) surviving mice. No residual disease was detected. FIG. 26A is a graph of bone marrow analysis for a Raji control mouse. FIG. 26B is a graph of bone marrow analysis for MATCH mouse 1. FIG. 26C is a graph of bone marrow analysis for MATCH mouse 2. FIG. 26D is a graph of bone marrow analysis for MATCH mouse 3. [00038] FIGS. 27A-B are graphs of In vivo evaluation of T cell-to-target cell ratio for a single 60 pg CD20-directed MATCH dose. NSG mice survival curve. Untreated control mice (orange) saw onset of hind-limb paralysis as soon as at Day 14. All untreated mice reached end-point by Day 17. Treated mice significantly increased survival over untreated with 5:1 T cell-to-target cell inoculated mice (blue) surviving the longest. FIG. 27A is a schematic of the dose schedule for the MATCH dosing of NSG mice. FIG. 27B is a graph showing mouse survival over time of varying treatment methods.

[00039] FIGS. 28A, 28B, 28C, 28D, and 28E are in vivo images taken weekly beginning 7 days post-inoculation. Mice were given an IP injection of 100 pL (30 mg mU¹) luciferin in PBS. After 15 min, mice were anesthetized and scanned for bioluminescence. All images are adjusted to the luminescence scale depicted to the right of each panel. FIG. 28A is an image of the mice in the control group. FIG. 28B is an image of the mice inoculated with a 1-to-1 T:B ratio. FIG. 28C is an image of the mice inoculated with a 2-to-1 T:B ratio. FIG. 28D is an image of the mice inoculated with a 5-to-1 T:B ratio. FIG. 28E is an image of the mice inoculated with a 10-to-1 T:B ratio.

[00040] FIGS. 29A, 29B, 29C, 29D, and 29E are graphs of the individual mouse body weights over time for all treatment groups. FIG. 29A is a graph of the mice treated with Raji control (saline). FIG. 29B is a graph of the mice treated with 2:1 T Cell-to-Raji. FIG. 29C is a graph of the mice treated with 1 :1 T Cell-to-Raji. FIG. 29D is a graph of the mice treated with 5:1 T Cell-to-Raji. FIG. 29E is a graph of the mice treated with 10:1 T Cell-to-Raji.

[00041] FIG. 30 is a graph of ex vivo bone marrow analysis for residual human CD3(+) leukocytes residing in sacrificed mouse bone marrow. Residual human T-cells were observed in all mice that received T-cell inoculation. Representative flow cytometry histograms overlays of CD3(+) cells from one mouse per group are shown.

[00042] FIGS. 31 A and 31 B are graphs of survival curve and dose schedule of an in vivo dose escalation of T-cell engager, Fab’cD3-MORF2 in a human xenograft NHL model. C.B-17 SCID mice were inoculated with a co-culture of Raji B-cells and healthy donor, naive T-cells in a 5:1 T cell-to-Raji cell ratio on Day 0. 1 h post-inoculation two-step CD20-directed MATCH was initiated. First the B-cell engager, Fab’_CD2o-MORF1 was administered via tail vein injection at a dose of 60 pg per mouse. 5 h later, the T-cell engager was administered via tail vein at various dose levels for each cohort of mice: 60 pg, 20 pg, 6 pg, and 2 pg. Mice were monitored with IVIS imaging and for hind-limb paralysis for experiment endpoint. FIG. 31 A is a schematic of the dose schedule of the in vivo dosing in C.B-17 SCID mice. FIG. 31 B is a graph of the survival curve of the mice given the in vivo dosing.

[00043] FIGS. 32A, 32B, 32C, 32D, 32E, 32F, and 32G show IVIS imaging of T-cell engager dose escalation in vivo experiment over the course of 13 weeks. FIG. 32A is an image of the mice in the control group. FIG. 32B is an image of the mice in the blinatumomab group. FIG. 32C is an image of the mice in the premixture group. FIG. 32D is an image of the mice in the 60 pg Fab’cD3-MORF2 group. FIG. 32E is an image of the mice in the 20 pg Fab’cD3-MORF2 group. FIG. 32F is an image of the mice in the 6 pg Fab’cD3-MORF2 group. FIG. 32G is an image of the mice in the 2 pg Fab’cD3-MORF2 group.

[00044] FIGS. 33A, 33B, 33C, 33D, 33E, 33F, and 33G are graphs of mouse weights for the T-cell engager dose escalation in vivo study. FIG. 33A is a graph of the mouse weights for the untreated group. FIG. 33B is a graph of the mouse weights for the 60 pg blinatumomab group. FIG. 33C is a graph of the mouse weights for the 60 pg premix group. FIG. 33D is a graph of the mouse weights for the 60 pg consecutive group. FIG. 33E is a graph of the mouse weights for the 20 pg consecutive group. FIG. 33F is a graph of the mouse weights for the 6 pg consecutive group. FIG. 33G is a graph of the mouse weights for the 2 pg consecutive group.

DETAILED DESCRIPTION

[00045] Described herein are kits comprising: (i) a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; (ii) a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of a target B-cell; and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell. The foregoing is termed “Multi-Antigen T-Cell Hybridizers” or “MATCH”.

[00046] MATCH extrapolate upon bispecific T-cell engaging technologies by reengineering the process of creating bispecific antibody molecules. Conceptually, MATCH expands the Drug-Free Macromolecular Therapeutics (DFMT) approach where receptor crosslinking and apoptosis initiation is mediated by hybridization of complementary morpholino oligonucleotides (MORFs), one bound to an antibody fragment, the other in multiple copies to a synthetic macromolecule or human serum albumin. In the MATCH technology hybridization of complementary MORFs mediates T-cell recruitment and activation against malignant B-cells. MATCH involves the creation of a cancer cell targeting motif library, based on antibody binding fragments (Fab’s), that have been modified to dimerize with T cell-engaging Fab’s. The cancer cell Fab’s are functionalized with a single stranded, 25 base-pair, MORF1. The T-cell Fab’ is functionalized with the complementary, single stranded 25 base-pair, MORF2. Complementary nucleotide strands allow for rapid and stable heterodimerization with high fidelity. Selfassembled MATCH conjugates resemble bispecific T-cell engaging constructs; however, the two-component nature of MATCH enables a modular and customizable approach to designing bi- and multi-specific T-cell recruitment therapies. MATCH’S “spl it-antibody”-like approach enables cancer cell-specific T-cell activation by matching a cancer cell’s unique surface protein expression profile with a corresponding MATCH targeting motif cocktail that can be customized from a library of targeting motifs.

[00047] MATCH is a two-component technology where two complementary morpholino oligomer nucleotides, “MORF1” and “MORF2”, are conjugated with various Fab’s - MORF1 to cancer antigen Fab’s, and MORF2 to a Fab’ that targets an antigen on the surface of an effector cells. One Fab’B-Cell-MORF1 species complements and dimerizes with one Fab’effector-cell- MORF2 molecule. The two-component T-cell activating system allows for interchangeability of target cell antigen engager, thus, T-cell activation can be achieved based on a cancer cell’s unique antigen expression profile by producing cell-specific MATCH therapies; and the dose of the T-cell engaging motif can be optimized independently from the target cell binding dose, thus, T-cell activation can be adjusted based on variations in T-cell counts and T cell-to-target cell ratio.

1. Definitions

[00048] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. The meaning and scope of the terms should be clear. In case of conflict, the present document, including definitions, take precedent over any dictionary or extrinsic definition. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular. Preferred methods and materials are described below, although methods and materials similar or equivalent to those described herein can be used in practice or testing of the present invention. All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. The materials, methods, and examples disclosed herein are illustrative only and not intended to be limiting.

[00049] The terms “comprise(s),” “include(s),” “having,” “has,” “can,” “contain(s),” and variants thereof, as used herein, are intended to be open-ended transitional phrases, terms, or words that do not preclude the possibility of additional acts or structures. The singular forms “a,” “and,” and “the” include plural references unless the context clearly dictates otherwise. The present disclosure also contemplates other embodiments “comprising,” “consisting of,” and “consisting essentially of,” the embodiments or elements presented herein, whether explicitly set forth or not.

[00050] For the recitation of numeric ranges herein, each intervening number there between with the same degree of precision is explicitly contemplated. For example, for the range of 6-9, the numbers 7 and 8 are contemplated in addition to 6 and 9, and for the range 6.0-7.0, the number 6.0, 6.1 , 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, and 7.0 are explicitly contemplated.

[00051] The term “about” or “approximately” as used herein as applied to one or more values of interest, refers to a value that is similar to a stated reference value, or within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, such as the limitations of the measurement system. In certain aspects, the term “about” refers to a range of values that fall within 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11 %, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value). Alternatively, “about” can mean within 3 or more than 3 standard deviations, per the practice in the art. Alternatively, such as with respect to biological systems or processes, the term “about” can mean within an order of magnitude, preferably within 5-fold, and more preferably within 2-fold, of a value.

[00052] As used herein, the term “analog” refers to a compound having a structure derived from the structure of a parent compound (e.g., a compound disclosed herein) and whose structure is sufficiently similar to those disclosed herein and based upon that similarity, would be expected by one skilled in the art to exhibit the same or similar activities and utilities as the claimed compounds, or to induce, as a precursor, the same or similar activities and utilities as the claimed compounds.

[00053] “Complement” or “complementary” as used herein means a nucleic acid can mean Watson-Crick (e.g., A-T/U and C-G) or Hoogsteen base pairing between nucleotides or nucleotide analogs of nucleic acid molecules. “Complementarity” refers to a property shared between two nucleic acid sequences, such that when they are aligned antiparallel to each other, the nucleotide bases at each position will be complementary. “Percent complementary” or “#% complementary”, with “#” representing a number, as used herein means the percentage of nucleotides that are complementary to a specific sequence. For example, a first morpholino oligonucleotide in which 18 of 20 nucleotides of the first morpholino oligonucleotide are complementary to a second morpholino oligonucleotide that specifically hybridizes with the second morpholino oligonucleotide, would be 90% complementary to the second morpholino oligonucleotide. In that example, the remaining noncomplementary nucleotides may be clustered or interspersed with complementary nucleotides and need not be contiguous to each other or to complementary nucleotides. Percent complementarity between particular stretches of nucleic acid sequences within nucleic acids can be determined routinely using BLAST programs (basic local alignment search tools) and PowerBLAST programs known in the art (Altschul et al., J. Mol. Biol., 1990, 215, 403-410; Zhang and Madden, Genome Res., 1997, 7, 649-656) or by using the Gap program (Wisconsin Sequence Analysis Package, Version 8 for Unix, Genetics Computer Group, University Research Park, Madison Wis.), using default settings, which uses the algorithm of Smith and Waterman (Adv. Appt. Math., 1981 , 2, 482-489).

[00054] The term “contacting” as used herein refers to bringing a disclosed composition, compound, kit, or conjugate together with an intended target (such as, e.g., a cell or population of cells, a receptor, an antigen, or other biological entity) in such a manner that the disclosed composition, compound, kit, or conjugate can affect the activity of the intended target (e.g., receptor, transcription factor, cell, population of cells, etc.), either directly (i.e., by interacting with the target itself), or indirectly (i.e., by interacting with another molecule, cell, co-factor, factor, or protein on which the activity of the target is dependent). In an aspect, a cell or population of cells, such as B-cells, can be contacted with a disclosed kit or conjugate.

[00055] The terms “control,” “reference level,” and “reference” are used herein interchangeably. The reference level may be a predetermined value or range, which is employed as a benchmark against which to assess the measured result. “Control group” as used herein refers to a group of control subjects. The predetermined level may be a cutoff value from a control group. The predetermined level may be an average from a control group. Cutoff values (or predetermined cutoff values) may be determined by Adaptive Index Model (AIM) methodology. Cutoff values (or predetermined cutoff values) may be determined by a receiver operating curve (ROC) analysis from biological samples of the patient group. ROC analysis, as generally known in the biological arts, is a determination of the ability of a test to discriminate one condition from another, e.g., to determine the performance of each marker in identifying a patient having CRC. A description of ROC analysis is provided in P.J. Heagerty et al. (Biometrics 2000, 56, 337-44), the disclosure of which is hereby incorporated by reference in its entirety. Alternatively, cutoff values may be determined by a quartile analysis of biological samples of a patient group. For example, a cutoff value may be determined by selecting a value that corresponds to any value in the 25th-75th percentile range, preferably a value that corresponds to the 25th percentile, the 50th percentile or the 75th percentile, and more preferably the 75th percentile. Such statistical analyses may be performed using any method known in the art and can be implemented through any number of commercially available software packages (e.g., from Analyse-it Software Ltd., Leeds, UK; StataCorp LP, College Station, TX; SAS Institute Inc., Cary, NC.). The healthy or normal levels or ranges for a target may be defined in accordance with standard practice. A control may be a subject or cell without a conjugate as detailed herein. A control may be a subject, or a sample therefrom, whose disease state is known. The subject, or sample therefrom, may be healthy, diseased, diseased prior to treatment, diseased during treatment, or diseased after treatment, or a combination thereof.

[00056] As used herein, the term “diagnosed” means having been subjected to a physical examination by a person of skill, for example, a physician, and found to have a condition that can be diagnosed or treated by the compounds, kits, or methods disclosed herein. For example, “diagnosed with a B-cell cancer” means having been subjected to a physical examination by a person of skill, for example, a physician, and found to have a condition that can be diagnosed or can be treated by a compound or kit that can prevent or inhibit malignant cell growth and/or induce apoptosis in a population of cells, such as B-cells. As a further example, “diagnosed with a need for inducing apoptosis” refers to having been subjected to a physical examination by a person of skill, for example, a physician, and found to have a condition characterized by malignant cell growth or other disease wherein inducing apoptosis of a population of cells would be beneficial to the subject. Such a diagnosis can be in reference to a disorder, such as cancer and an autoimmune disorder, and the like, as discussed herein.

[00057] “Identical” or “identity” as used herein in the context of two or more polynucleotide or polypeptide sequences means that the sequences have a specified percentage of residues that are the same over a specified region. The percentage may be calculated by optimally aligning the two sequences, comparing the two sequences over the specified region, determining the number of positions at which the identical residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the specified region, and multiplying the result by 100 to yield the percentage of sequence identity. In cases where the two sequences are of different lengths or the alignment produces one or more staggered ends and the specified region of comparison includes only a single sequence, the residues of single sequence are included in the denominator but not the numerator of the calculation. When comparing DNA and RNA, thymine (T) and uracil (U) may be considered equivalent. Identity may be performed manually or by using a computer sequence algorithm such as BLAST or BLAST 2.0.

[00058] As used herein, “homolog” or “homologue” refers to a polypeptide or nucleic acid with homology to a specific known sequence. Specifically disclosed are variants of the nucleic acids and polypeptides herein disclosed which have at least 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, 49, 50, 51 , 52, 53, 54, 55, 56, 57, 58, 59, 60, 61 , 62, 63, 64, 65, 66, 67, 68, 69, 70, 71 , 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, or more percent homology to the stated or known sequence. Those of skill in the art readily understand how to determine the homology of two or more proteins or two or more nucleic acids. For example, the homology can be calculated after aligning the two or more sequences so that the homology is at its highest level. It is understood that one way to define any variants, modifications, or derivatives of the disclosed genes and proteins herein is through defining the variants, modification, and derivatives in terms of homology to specific known sequences.

[00059] “Natural” as used herein in refers to a molecule or subject found in nature that is unaltered from the form found in nature. For example, natural DNA is an organic chemical that contains genetic information and instructions for protein synthesis that is found in most cells of every organism. [00060] “Normal” as used herein refers to cells that can reproduce when and where they need to, stick together in the right place in the body, and self-destruct when they become damaged or too old. “Normal” as used herein also refers to wildtype animals that are healthy (no diseases or disorders) and are unmodified (e.g., not genetically modified).

[00061] “Nucleic acid” or “oligonucleotide” or “polynucleotide” as used herein means at least two nucleotides covalently linked together. The depiction of a single strand also defines the sequence of the complementary strand. Thus, a polynucleotide also encompasses the complementary strand of a depicted single strand. Many variants of a polynucleotide may be used for the same purpose as a given polynucleotide. Thus, a polynucleotide also encompasses substantially identical polynucleotides and complements thereof. A single strand provides a probe that may hybridize to a target sequence under stringent hybridization conditions. Thus, a polynucleotide also encompasses a probe that hybridizes under stringent hybridization conditions. Polynucleotides may be single stranded or double stranded or may contain portions of both double stranded and single stranded sequence. The polynucleotide can be nucleic acid, natural or synthetic, DNA, genomic DNA, cDNA, RNA, or a hybrid, where the polynucleotide can contain combinations of deoxyribo- and ribo-nucleotides, and combinations of bases including, for example, uracil, adenine, thymine, cytosine, guanine, inosine, xanthine hypoxanthine, isocytosine, and isoguanine. Polynucleotides can be obtained by chemical synthesis methods or by recombinant methods.

[00062] A “peptide” or “polypeptide” is a linked sequence of two or more amino acids linked by peptide bonds. The polypeptide can be natural, synthetic, or a modification or combination of natural and synthetic. Peptides and polypeptides include proteins such as binding proteins, receptors, and antibodies. The terms “polypeptide”, “protein,” and “peptide” are used interchangeably herein. “Primary structure” refers to the amino acid sequence of a particular peptide. “Secondary structure” refers to locally ordered, three dimensional structures within a polypeptide. These structures are commonly known as domains, for example, enzymatic domains, extracellular domains, transmembrane domains, pore domains, and cytoplasmic tail domains. “Domains” are portions of a polypeptide that form a compact unit of the polypeptide and are typically 15 to 350 amino acids long. Exemplary domains include domains with enzymatic activity or ligand binding activity. Typical domains are made up of sections of lesser organization such as stretches of beta-sheet and alpha-helices. “Tertiary structure” refers to the complete three-dimensional structure of a polypeptide monomer. “Quaternary structure” refers to the three-dimensional structure formed by the noncovalent association of independent tertiary units. A “motif” is a portion of a polypeptide sequence and includes at least two amino acids. A motif may be 2 to 20, 2 to 15, or 2 to 10 amino acids in length. In some embodiments, a motif includes 3, 4, 5, 6, or 7 sequential amino acids. A domain may be comprised of a series of the same type of motif.

[00063] “Sample” or “test sample” as used herein can mean any sample in which the presence and/or level of an antigen is to be detected or determined or any sample comprising a kit as detailed herein. Samples may include liquids, solutions, emulsions, or suspensions. Samples may include a culture sample, such as a cell line. Samples may include a medical sample. Samples may include any biological fluid or tissue, such as blood, whole blood, fractions of blood such as plasma and serum, muscle, interstitial fluid, sweat, saliva, urine, tears, synovial fluid, bone marrow, cerebrospinal fluid, nasal secretions, sputum, amniotic fluid, bronchoalveolar lavage fluid, gastric lavage, emesis, fecal matter, lung tissue, peripheral blood mononuclear cells, total white blood cells, lymph node cells, spleen cells, tonsil cells, cancer cells, tumor cells, bile, digestive fluid, skin, or combinations thereof. In some embodiments, the sample comprises an aliquot. In other embodiments, the sample comprises a biological fluid. Samples can be obtained by any means known in the art. The sample can be used directly as obtained from a patient or can be pre-treated, such as by filtration, distillation, extraction, concentration, centrifugation, inactivation of interfering components, addition of reagents, and the like, to modify the character of the sample in some manner as discussed herein or otherwise as is known in the art.

[00064] “Subject” and “patient” as used herein interchangeably refers to any vertebrate, including, but not limited to, a mammal that wants or is in need of the herein described kits or methods. The subject may be a human or a non-human. The subject may be a vertebrate. The subject may be a mammal. The mammal may be a primate or a non-primate. The mammal can be a non-primate such as, for example, cow, pig, camel, llama, hedgehog, anteater, platypus, elephant, alpaca, horse, goat, rabbit, sheep, hamsters, guinea pig, cat, dog, rat, and mouse. The mammal can be a primate such as a human. The mammal can be a non- human primate such as, for example, monkey, cynomolgus monkey, rhesus monkey, chimpanzee, gorilla, orangutan, and gibbon. The subject may be of any age or stage of development, such as, for example, an adult, an adolescent, or an infant. The subject may be male. The subject may be female. In some embodiments, the subject has cancer or an autoimmune disorder. The subject may be undergoing other forms of treatment. [00065] A “patient” or a “subject in need thereof” refers to a subject afflicted with one or more diseases or disorders, such as a B-cell malignancy, an inflammatory disorder, and an autoimmune disease with B-cell involvement. A patient may have been diagnosed with a need for treatment of one or more of the aforementioned diseases or disorders prior. A patient may have been diagnosed with a need for inducing apoptosis of malignant cells, such as, for example, malignant B-cells.

[00066] “Substantially identical” can mean that a first and second amino acid or polynucleotide sequence are at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% over a region of 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100 amino acids or nucleotides, respectively.

[00067] “Target” as used herein refers to any peptide or polypeptide of a known or putative protein product. The target may be an antigen on the surface of a cell that is involved in a disease or disorder such as cancer or an autoimmune disease.

[00068] As used herein, a “targeting moiety” can be specific to a recognition molecule on the surface of a cell or a population of cells, such as, for example B-cells. In an aspect of the disclosed kits and methods, a targeting moiety can include, but is not limited to: a monoclonal antibody, a polyclonal antibody, full-length antibody, a chimeric antibody, Fab’, Fab, F(ab)2, F(ab’)2, a single domain antibody (DAB), Fv, a single chain Fv (scFv), a minibody, a diabody, a triabody, hybrid fragments, a phage display antibody, a ribosome display antibody, a peptide, a peptide ligand, a hormone, a growth factor, or a cytokine.

[00069] “Treatment” or “treating” or “treatment” when referring to protection of a subject from a disease, means suppressing, repressing, reversing, alleviating, ameliorating, or inhibiting the progress of disease, or completely eliminating a disease. A treatment may be either performed in an acute or chronic way. The term also refers to reducing the severity of a disease or symptoms associated with such disease prior to affliction with the disease. Preventing the disease involves administering at least one component of the presently disclosed kit to a subject prior to onset of the disease. Suppressing the disease involves administering at least one component of the presently disclosed kit to a subject after induction of the disease but before its clinical appearance. Repressing or ameliorating the disease involves administering at least one component of the presently disclosed kit to a subject after clinical appearance of the disease. [00070] “Variant” used herein with respect to a polynucleotide means (i) a portion or fragment of a referenced nucleotide sequence; (ii) the complement of a referenced nucleotide sequence or portion thereof; (iii) a nucleic acid that is substantially identical to a referenced nucleic acid or the complement thereof; or (iv) a nucleic acid that hybridizes under stringent conditions to the referenced nucleic acid, complement thereof, or a sequences substantially identical thereto.

[00071] “Variant” with respect to a peptide or polypeptide that differs in amino acid sequence by the insertion, deletion, or conservative substitution of amino acids, but retain at least one biological activity. Variant may also mean a protein with an amino acid sequence that is substantially identical to a referenced protein with an amino acid sequence that retains at least one biological activity. Representative examples of “biological activity” include the ability to be bound by a specific antibody or polypeptide or to promote an immune response. Variant can mean a functional fragment thereof. Variant can also mean multiple copies of a polypeptide. The multiple copies can be in tandem or separated by a linker. A conservative substitution of an amino acid, for example, replacing an amino acid with a different amino acid of similar properties (for example, hydrophilicity, degree and distribution of charged regions) is recognized in the art as typically involving a minor change. These minor changes may be identified, in part, by considering the hydropathic index of amino acids, as understood in the art (Kyte et al., J.

Mol. Biol. 1982, 157, 105-132). The hydropathic index of an amino acid is based on a consideration of its hydrophobicity and charge. It is known in the art that amino acids of similar hydropathic indexes may be substituted and still retain protein function. In one aspect, amino acids having hydropathic indexes of ±2 are substituted. The hydrophilicity of amino acids may also be used to reveal substitutions that would result in proteins retaining biological function. A consideration of the hydrophilicity of amino acids in the context of a peptide permits calculation of the greatest local average hydrophilicity of that peptide. Substitutions may be performed with amino acids having hydrophilicity values within ±2 of each other. Both the hydrophobicity index and the hydrophilicity value of amino acids are influenced by the particular side chain of that amino acid. Consistent with that observation, amino acid substitutions that are compatible with biological function are understood to depend on the relative similarity of the amino acids, and particularly the side chains of those amino acids, as revealed by the hydrophobicity, hydrophilicity, charge, size, and other properties.

2. Conjugates [00072] Provided herein are conjugates comprising a first targeting moiety and a first morpholino oligonucleotide. The first targeting moiety may be adapted to specifically bind to a first antigen on the surface of a target effector cell.

[00073] Also provided herein is a plurality of additional conjugates. The additional conjugates may include a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The second targeting moiety may be adapted to bind to a second antigen that is on the surface of a target B-cell.

[00074] The additional conjugates may also include a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The third targeting moiety may be adapted to bind to a third antigen that is on the surface of the target B-cell.

[00075] The additional conjugates may also include a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The fourth targeting moiety may be adapted to bind to a fourth antigen that is on the surface of the target B-cell.

[00076] The additional conjugates may also include a fifth conjugate comprising a fifth targeting moiety and a fifth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The fifth targeting moiety may be adapted to bind to a fifth antigen that is on the surface of the target B-cell.

[00077] The additional conjugates may also include a sixth conjugate comprising a sixth targeting moiety and a sixth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The sixth targeting moiety may be adapted to bind to a sixth antigen that is on the surface of the target B-cell.

[00078] The additional conjugates may also include a seventh conjugate comprising a seventh targeting moiety and a seventh morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The seventh targeting moiety may be adapted to bind to a seventh antigen that is on the surface of the target B-cell.

[00079] The additional conjugates may also include an eighth conjugate comprising an eighth targeting moiety and an eighth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The eighth targeting moiety may be adapted to bind to an eighth antigen that is on the surface of the target B-cell.

[00080] The additional conjugates may also include a ninth conjugate comprising a ninth targeting moiety and a ninth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The ninth targeting moiety may be adapted to bind to a ninth antigen that is on the surface of the target B-cell.

[00081] The additional conjugates may also include a tenth conjugate comprising a tenth targeting moiety and a tenth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide. The tenth targeting moiety may be adapted to bind to a tenth antigen that is on the surface of the target B-cell.

[00082] Types of conjugation and methods for conjugating are known to the art. A targeting moiety of a disclosed conjugate can be conjugated to a morpholino oligonucleotide via, for example, a covalent bond, a thiol group, a thioether bond, a thiol-maleimide bond, a thiol- vinylsulfone bond, a thiol-halogeno bond, a thiol-pentafluorophenyl ester bond, a thiol-ene bond, or a thiol-yne bond. A targeting moiety of a disclosed conjugate can be coupled to a morpholino oligonucleotide by a linker.

[00083] For example, the first targeting moiety may be coupled to the first morpholino oligonucleotide by a first linker; the second targeting moiety may be coupled to the second morpholino by a second linker; the third targeting moiety may be coupled to the third morpholino oligonucleotide by a third linker; the fourth targeting moiety may be coupled to the fourth morpholino by a fourth linker; the fifth targeting moiety may be coupled to the fifth morpholino oligonucleotide by a fifth linker; the sixth targeting moiety may be coupled to the sixth morpholino by a sixth linker; the seventh targeting moiety may be coupled to the seventh morpholino oligonucleotide by a seventh linker; the eighth targeting moiety may be coupled to the eighth morpholino by an eighth linker; the ninth targeting moiety may be coupled to the ninth morpholino by a ninth linker; the tenth targeting moiety may be coupled to the tenth morpholino by a tenth linker.

[00084] A linker may be from about 10 A to about 100 A, about 15 A to about 100 A, about 20 A to about 100 A, about 25 A to about 100 A, about 30 A to about 100 A, about 35 A to about 100 A, about 40 A to about 100 A, about 45 A to about 100 A, about 50 A to about 100 A, about 55 A to about 100 A, about 60 A to about 100 A, about 65 A to about 100 A, about 70 A to about 100 A, about 75 A to about 100 A, about 80 A to about 100 A, about 85 A to about 100 A, about 90 A to about 100 A, about 95 A to about 100 A, about 10 A to about 95 A, about 10 A to about 90 A, about 10 A to about 85 A, about 10 A to about 80 A, about 10 A to about 75 A, about 10 A to about 70 A, about 10 A to about 65 A, about 10 A to about 60 A, about 10 A to about 55 A, about 10 A to about 50 A, about 10 A to about 45 A, about 10 A to about 40 A, about 10 A to about 35 A, about 10 A to about 30 A, about 10 A to about 25 A, about 10 A to about 20 A, or about 10 A to about 15 A in length.

[00085] A linker may comprise polyethylene glycol (PEG) or succinimidyl-4-(N- maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC). The first linker, the second linker, the third linker, the fourth linker, the fifth linker, the sixth linker, the seventh linker, the eighth linker, the ninth linker, and the tenth linker may each independently have either the same or a different molecular weight relative to one another.

[00086] A conjugate as described herein may comprise a detectable label. Detectable labels are known to one of skill in the art and include, but are not limited to: rhodamine, FITC, Cy3, Cy3.5, Cy5, Texas Red, Alexa Fluor 488, Alexa Fluor 610, Alexa Fluor 647, and Alexa Fluor 750.

[00087] A plurality of additional conjugates as described herein may comprise one or more morpholinos, where each morpholino is coupled to a targeting moiety. The one or more morpholines can comprise 1 morpholino, 2 morpholinos, 3 morpholinos, 4 morpholinos, 5 morpholinos, 6 morpholinos, 7 morpholinos, 8 morpholinos, 9 morpholinos, or 10 morpholinos. When the plurality of additional conjugates comprises more than one morpholino, the morpholinos can have the same sequence, can have different sequences, or a combination thereof. When the plurality of additional conjugates comprises more than one morpholino, the targeting moieties can be adapted to bind to the same antigen, different antigens, or a combination thereof. a. Morpholino Oligonucleotide

[00088] A biocompatible, synthetic oligonucleotide analogue with a chemically modified backbone may be used. The conjugates disclosed herein may comprise a biocompatible oligonucleotide. The conjugates disclosed herein may comprise a non-degradable oligonucleotide. The conjugates disclosed herein may comprise a water-soluble oligonucleotide. The conjugates disclosed herein may comprise a charge-neutral oligonucleotide. The conjugates disclosed herein may comprise a biocompatible and non- degradable oligonucleotide. The conjugates disclosed herein may comprise a water-soluble and charge-neutral oligonucleotide. The conjugates disclosed herein may comprise an oligonucleotide that has one or more of the following properties: biocompatible, non-degradable, water-soluble, and charge neutral. For example, an oligonucleotide can be biocompatible, non- degradable, water-soluble, and charge neutral.

[00089] An oligonucleotide can be a peptide nucleic acid. An oligonucleotide can be a morpholino. As used herein, “morpholino oligonucleotide” and “morpholino” are used interchangeably. A morpholino as described herein may not bind to any mRNA target of a genome, such as the human genome or the mouse genome. A morpholino as described herein may not be self-complementary. A morpholino as described herein can be succinimidyl-4-(N- maleimidomethyl)cyclohexane-1-carboxylate (SMCC) modified. A morpholino as described herein may be amine-derivatized. Derivatization, which typically involves the addition of a nucleophile as a functional group, and which includes amine derivatization and thiol derivatization, is known to the art. A morpholino as described herein may be generated through the use of an amine-pentafluorophenyl ester, an amine-succinimidoxy ester, or an aminecarboxyl. A morpholino as described herein may be thiol-derivatized. A morpholino as described herein may be generated through the use of thiol-maleimide.

[00090] The properties of several analogues compared with those of natural DNA is as follows: natural DNA is degradable, anionic, and soluble; mDNA is resistant, neutral, and nonsoluble; S-DNA is resistant, anionic, and soluble; morpholinos are resistant, neutral, and soluble; and PNA is resistant, neutral, and non-soluble. Based on these properties, the disclosed kits and methods are not compatible with natural DNA or RNA. Rather, as the analogue must be biocompatible and non-degradable, the disclosed kits and methods can utilize phophorodiamidate morpholino oligonucleotides (referred to as “morpholino oligonucleotides”, “morpholinos”, or “MORFs” herein). Morpholinos have a chemically modified, non-charged backbone and are assembled from four different subunits, each of which contains one of the four nucleobases (A, T, G, and C) linked to a 6-membered morpholine ring. The subunits are joined by non-ionic phosphordiamidate linkages to generate a morpholino oligonucleotide. Morpholinos also possess strong binding affinity (i.e. , Kd from the low nM to pM levels), high sequence specificity, and well-demonstrated safety profiles. Furthermore, the immunogenicity of morpholinos is highly sequence dependent, and therefore, can be addressed. The synthesis, structures, and binding characteristics of morpholinos are detailed in U.S. Patent Nos. 5,698,685; 5,217,866; 5,142,047; 5,034,506; 5,166,315; 5,521 ,063; and 5,506,337, each of which is incorporated herein by reference in its entirety.

[00091] A disclosed morpholino having a longer length provides a higher specificity and a stronger binding affinity. However, such morpholinos also have poorer water-solubility. In the art, a 14-base pair (bp) to 15-bp morpholino is considered the minimal length necessary to maintain ideal targeting effects. A 25-bp morpholino can ensure strong binding affinity and good water-solubility (about 5-30 mM). For example, using 25 bp morpholinos in the disclosed kits and methods can avoid the impact of steric hindrance on the hybridization of the MORFIs and MORF2s disclosed in TABLE 1. A longer sequence can provide better “steric flexibility” for hybridization. Accordingly, in the compositions and methods disclosed herein, morpholinos can comprise from about 10 nucleotides to about 30 nucleotides. A morpholino can be from about 10 nucleotides to about 30 nucleotides, about 12 nucleotides to about 30 nucleotides, about 14 nucleotides to about 30 nucleotides, about 16 nucleotides to about 30 nucleotides, about 18 nucleotides to about 30 nucleotides, about 20 nucleotides to about 30 nucleotides, about 22 nucleotides to about 30 nucleotides, about 24 nucleotides to about 30 nucleotides, about 26 nucleotides to about 30 nucleotides, about 28 nucleotides to about 30 nucleotides, about 10 nucleotides to about 28 nucleotides, about 10 nucleotides to about 26 nucleotides, about 10 nucleotides to about 24 nucleotides, about 10 nucleotides to about 22 nucleotides, about 10 nucleotides to about 20 nucleotides, about 10 nucleotides to about 18 nucleotides, about 10 nucleotides to about 16 nucleotides, about 10 nucleotides to about 14 nucleotides, or about 10 nucleotides to about 12 nucleotides in length. A morpholino can be 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides in length.

[00092] The A/T/C/G content of a disclosed morpholino oligonucleotide can be determined based on three factors: G+C content (number or percent of G’s and C’s); G content (number or percent of G’s); and C content (number or percent of C’s). Regarding G+C content, a disclosed morpholino can comprise a G+C content of from about 35% to about 65%. This range can provide optimal binding efficacy and specificity. Regarding G content, a disclosed morpholino can comprise a G content of less than about 36%. This level of G content can provide good aqueous solubility. However, repeats of 4 or more G’s should be avoided. Regarding C content, a disclosed morpholino can comprise a C content of less than about 7. This level of C content can ensure that the unfavorable effect of enhancing kidney accumulation of a morpholino can be avoided. Liu et al. shows kidney accumulation of morpholinos comprising varying levels of C content, where a morpholino having 25 C's had the highest percent accumulation in the kidneys of normal mice just 3 hours post-injection (Liu et al., Eur. J. Nucl.

Med. Mol. Imaging 31 (2004) 417-424).

[00093] For example, the first morpholino conjugated to the first targeting moiety as described herein can comprise more A’s and less C’s whereas the second, third, fourth, fifth, sixth, seventh, or eighth morpholinos conjugated to the second, third, fourth, fifth, sixth, seventh, or eighth targeting moiety as described herein can comprise more C’s and less A’s. Accordingly, a 25 base pair (bp) morpholino can comprise 3 C’s, 6 G’s, 12 A’s, and 4 T’s (G+C = 36%, G = 24%). A complementary 25 bp morpholino can comprise 6 C’s, 3 G’s, 4 A’s, and 12 T’s (G+C = 36%, G = 12%).

[00094] After the nucleobase composition of each morpholino is determined, a publicly accessible, online sequence “scrambler” can be used to ensure minimal off-target binding with human and murine mRNA (Chu et al., ACS Nano 8 (2014) 719-730). Furthermore, publicly accessible, online sequence analysis software can be used to ensure minimal selfcomplementarity (Chu et al.). For example, in the experiments disclosed in U.S. Patent No. 10,251 ,906 B2, which is incorporated herein by reference in its entirety, when performing sequence analysis to avoid self-complementarity, the “Minimum base pairs required for selfdimerization” and “Minimum base pairs required for a hairpin” were set to “2” and “2” (for 10 bp and 12 bp); “3” and “3” (for 15 bp, 18 bp, 20 bp, 23 bp, and 25 bp); “4” and “4” (for 28 bp, 30 bp, 32 bp, and 35 bp); and “5” and “4” (for 38 bp and 40 bp). TABLE 1 provides a listing of exemplary morpholinos for use in the disclosed kits and methods.

TABLE 1. Listing of Exemplary Morpholinos.

[00095] A morpholino may be SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11 , SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21 , SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31 , SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41 , SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51 , or SEQ ID NO: 52.

[00096] Hybridization between a pair of morpholines disclosed herein can be achieved by base-pairing (i.e., specific hydrogen bonding patterns). The hybridization can be maintained by base-stacking (i.e., pi interactions). It is noted that the hybridization between a pair of disclosed morpholinos is more specific that the formation of coiled-coil peptides.

[00097] The morpholinos utilized herein can be completely complementary (100%) or can be less than completely complementary. The percent complementarity of the morpholino of the first conjugate and each morpholino of the additional conjugates can be from about 80% to about 85%, about 85% to about 90%, about 90% to about 95%, or about 95% to about 100% complementary. The percent complementarity of the morpholino of the first conjugate and each morpholino of the additional conjugates can be about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% complementary. The percent complementarity of the morpholino of the first conjugate and each morpholino of the additional conjugates can be at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% complementary.

[00098] The morpholino of the first conjugate and each morpholino of the additional conjugates can have an equilibrium dissociation constant (Kd) of less than 15 nM. The morpholino of the first conjugate and each morpholino of the additional conjugates can have a binding constant (Kb) smaller than 10^-7 M. The morpholino of the first conjugate and each morpholino of the additional conjugates can have a binding constant (Kb) smaller than 10'⁹ M. b. Targeting Moiety

[00099] A targeting moiety can be specific for a non-internalizing cell surface molecule or slowly internalizing cell surface molecule. Examples of a non-internalizing cell surface molecule or a slowly internalizing cell surface molecule are known to one of skill in the art. A noninternalizing cell surface molecule can be a receptor. A slowly internalizing cell surface molecule can be a receptor. For example, non-internalizing cell surface molecules or slowly internalizing cell surface molecules include, but are not limited to: CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38, CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, programmed death-ligand 1 (PD-L1), programmed death protein 1 (PD-1), B-cell maturation antigen (BCMA), signaling lymphocyte activation molecular family 7 (SLAMF7; i.e. CS-1 or CD319), G protein-coupled receptor class C group 5 member D (GPRC5D), CD3, CD16, CD14, and CD64.

[000100] A non-internalizing cell surface molecule or slowly internalizing cell surface molecule can be on a cell or a population of cells. A cell or a population of cells can be target cells such as B-cells. The B-cells can be normal B-cells. The B-cells can be malignant B-cells. A cell or a population of cells can be effector cells including, but not limited to, T-cells, natural killer cells (NK cells), and macrophages. The T-cells, NK cells, and macrophages can be normal T-cells, NK cells, and macrophages.

[000101] A targeting moiety may be a polysaccharide, a peptide ligand, an aptamer, an antibody, a Fab’ fragment, or a single-chain variable fragment. A targeting moiety may be an antibody or a Fab’ fragment.

[000102] In an embodiment, the first targeting moiety may be a first antibody or a first Fab’ fragment of the first antibody. The first antigen may be CD3, CD16, CD14, or CD64. The first antibody may be anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), anti-CD14 (clone: UCHM-1), or anti-CD64 (clone 10.1)

[000103] In an embodiment, the second, third, fourth, fifth, sixth, seventh, eighth, ninth, or tenth targeting moieties may each be a second, third, fourth, fifth, sixth, seventh, eighth, ninth, or tenth antibody; or a second, third, fourth, fifth, sixth, seventh, eighth, ninth, or tenth Fab’ fragment of the second, third, fourth, fifth, sixth, seventh, eighth, ninth, or tenth antibody. The second, third, fourth, fifth, sixth, seventh, eighth, ninth, or tenth antigens may each independently be CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38, CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1, BCMA, SLAMF7 (CS-1), or GPRC5D. The second, third, fourth, fifth, sixth, seventh, eighth, ninth, or tenth antibodies may each independently be UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, or Talquetamab. c. Synthesis of Conjugates i) Synthesis of Conjugates Comprising a Targeting Moiety and a Morpholino Oligonucleotide

[000104] Disclosed herein are processes of synthesizing a conjugate comprising a targeting moiety and a morpholino oligonucleotide. The process may comprise obtaining a targeting moiety, modifying an oligonucleotide, and conjugating the targeting moiety with the oligonucleotide. A targeting moiety can be conjugated to the oligonucleotide via a thioether bond. An oligonucleotide can be SMCC modified. The oligonucleotide can contain a 3’- maleimido group. A process of synthesizing a conjugate can comprise introducing a detectable label. A targeting moiety can be a targeting moiety as described above. For example, a targeting moiety can be a Fab’ fragment specific for CD20. An oligonucleotide can be an oligonucleotide as described above. For example, an oligonucleotide can be a morpholino comprising from about 10 bp to about 30 bp. ii) Synthesis of Compositions Comprising Conjugates Comprising a Targeting Moiety and a Morpholino Oligonucleotide

[000105] Disclosed herein are processes of synthesizing a composition comprising conjugates comprising targeting moieties and morpholino oligonucleotides. The process may comprise contacting a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide with at least a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide. The first morpholino oligonucleotide of the first conjugate may hybridize to the second morpholino oligonucleotide of the second conjugate. For example, a first targeting moiety can be a Fab’ fragment specific for CD3, the second targeting moiety can be a Fab’ fragment specific for CD20, and each of the morpholino oligonucleotides can comprise from about 10 bp to about 30 bp, where the morpholino of the first conjugate is complementary to the morpholino of the second conjugate.

[000106] Alternatively, the process may comprise contacting a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide with at least a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide. The second morpholino oligonucleotide of the second conjugate may not hybridize to the third morpholino oligonucleotide of the third conjugate. In some instances, the sequence of the second morpholino oligonucleotide may be identical to the sequence of the third morpholino oligonucleotide. For example, a second targeting moiety can be a Fab’ fragment specific for CD20, the third targeting moiety can be a Fab’ fragment specific for CD5, and each of the morpholino oligonucleotides can comprise from about 10 bp to about 30 bp, where the morpholino of the second conjugate is identical to the morpholino of the third conjugate.

3. Pharmaceutical Compositions

[000107] Further provided herein are pharmaceutical compositions comprising one or more of the above-described conjugates. The conjugates described herein may be formulated separately or co-formulated. For example, a pharmaceutical composition may comprise a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide as described herein, and a pharmaceutically acceptable carrier. In another example, a pharmaceutical composition may comprise one conjugate of the plurality of additional conjugates as described herein and a pharmaceutically acceptable carrier. In another example, a pharmaceutical composition may comprise more than one of the plurality of additional conjugates as described herein and a pharmaceutically acceptable carrier. In another example, a pharmaceutical composition may comprise a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide as described herein, one or more of the plurality of additional conjugates as described herein, and a pharmaceutically acceptable carrier.

[000108] In some embodiments, the pharmaceutical composition may comprise about 1 ng to about 10 mg of a conjugate. One or more of the conjugates as detailed herein, or at least one component thereof, may be formulated into pharmaceutical compositions in accordance with standard techniques well known to those skilled in the pharmaceutical art. The pharmaceutical compositions can be formulated according to the mode of administration to be used. In cases where pharmaceutical compositions are injectable pharmaceutical compositions, they are sterile, pyrogen free, and particulate free. An isotonic formulation is preferably used. Generally, additives for isotonicity may include sodium chloride, dextrose, mannitol, sorbitol, and lactose. In some cases, isotonic solutions such as phosphate buffered saline are preferred. Stabilizers can include gelatin and albumin. A vasoconstriction agent may be added to the formulation.

[000109] The composition may further comprise a pharmaceutically acceptable excipient. The pharmaceutically acceptable excipient may be functional molecules such as vehicles, adjuvants, carriers, or diluents. The term “pharmaceutically acceptable carrier,” may be a non-toxic, inert solid, semi-solid or liquid filler, gas, diluent, encapsulating material or formulation auxiliary of any type. Pharmaceutically acceptable carriers include, for example, diluents, lubricants, binders, disintegrants, colorants, flavors, sweeteners, antioxidants, preservatives, glidants, solvents, suspending agents, wetting agents, surfactants, emollients, propellants, humectants, powders, pH adjusting agents, and combinations thereof.

[000110] Examples of solid carriers include, but are not limited to, lactose, terra alba, sucrose, talc, gelatin, agar, pectin, acacia, magnesium stearate, and stearic acid. Examples of liquid carriers include, but are not limited to, sugar syrup, peanut oil, olive oil, and water. Examples of gaseous carriers include, but are not limited to, carbon dioxide and nitrogen.

[000111] In preparing the conjugates described herein for oral dosage form, any convenient pharmaceutical media can be employed. For example, water, glycols, oils, alcohols, flavoring agents, preservatives, coloring agents, and the like can be used to form oral liquid preparations such as suspensions, elixirs, and solutions. Whereas carriers such as starches, sugars, microcrystalline cellulose, diluents, granulating agents, lubricants, binders, disintegrating agents, and the like can be used to form oral solid preparations such as powders, capsules, and tablets. Optionally, tablets can be coated by standard aqueous or nonaqueous techniques. A tablet containing a conjugate disclosed herein can be prepared by compression or molding, optionally with one or more accessory ingredients or adjuvants. Compressed tablets can be prepared by compressing, in a suitable machine, a disclosed complex of composition in a free- flowing form such as powder or granules, optionally mixed with a binder, lubricant, inert diluent, surface active or dispersing agent. Molded tablets can be made by molding in a suitable machine, a mixture of the powdered compound moistened with an inert liquid diluent. [000112] It is understood that the disclosed compositions can be prepared from the disclosed conjugates. It is also understood that the disclosed compositions can be employed in the disclosed kits and methods.

4. Kits

[000113] Also provided herein are kits, which may be used in the methods described herein. The kits comprise one or more conjugates or a composition comprising the same, as described above, and instructions for using said conjugates or composition. The kit may comprise a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide as described herein. The kit may comprise at least a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide as described herein and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide as described herein. The kit may comprise a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide and a plurality of additional conjugates that may comprise at least a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide.

[000114] The kit may comprise at least one morpholino comprising or having a polynucleotide sequence of SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11 , SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21 , SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31 , SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41 , SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51 , or SEQ ID NO: 52, a complement thereof, a variant thereof, or fragment thereof. The plurality of additional conjugates may comprise morpholines having the same polynucleotide sequence.

[000115] The plurality of additional conjugates may be co-formulated. The first conjugate and the plurality of additional conjugates may be co-formulated. The plurality of additional conjugates may be co-packaged. The first conjugate and the plurality of additional conjugates may be co-packaged. [000116] The kit may include instructions for using the conjugates. Instructions included in kits may be affixed to packaging material or may be included as a package insert. While the instructions are typically written on printed materials, they are not limited to such. Any medium capable of storing such instructions and communicating them to an end user is contemplated by this disclosure. Such media include, but are not limited to, electronic storage media (e.g., magnetic discs, tapes, cartridges, chips), optical media (e.g., CD ROM), and the like. As used herein, the term “instructions” may include the address of an internet site that provides the instructions.

5. Administration

[000117] The conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, may be administered to a subject. Such conjugates or compositions can be administered in dosages and by techniques well known to those skilled in the medical arts taking into consideration such factors as the age, sex, weight, and condition of the particular subject, and the route of administration. The presently disclosed conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, may be administered to a subject by different routes including orally, parenterally, sublingually, transdermally, rectally, transmucosally, topically, intranasal, intravaginal, via inhalation, via buccal administration, intrapleurally, intravenous, intraarterial, intraperitoneal, subcutaneous, intradermally, epidermally, intramuscular, intranasal, intrathecal, intracranial, and intraarticular or combinations thereof. In certain embodiments, the conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, are administered to a subject orally, intravenously, or a combination thereof. The conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, may be delivered to a subject by several technologies including injection, liposome mediated, or nanoparticle facilitated. For veterinary use, the conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, may be administered as a suitably acceptable formulation in accordance with normal veterinary practice. The veterinarian may readily determine the dosing regimen and route of administration that is most appropriate for a particular animal. The conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, may be administered by traditional syringes, needleless injection devices, or other physical methods. [000118] Upon delivery of the presently disclosed conjugates or kits as detailed herein, or at least one component thereof, or the pharmaceutical compositions comprising the same, and thereupon contacting the conjugate with the cells of the subject, surface molecules of the contacted cells bind the targeting moiety of the conjugates.

6. Methods a. Methods of Inducing Apoptosis

[000119] Provided herein are methods of inducing apoptosis of a target B-cell. The methods may include contacting a population of cells with a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; and contacting the population of cells with a plurality of additional conjugates. The plurality of additional conjugates may comprise a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of the target B-cell; and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell. The first morpholino oligonucleotide may hybridize with at least the second morpholino oligonucleotide, and the target effector cell induces apoptosis of the target B-cell.

[000120] The population of cells may be in a subject. The subject may be a mammalian subject having cancer or an autoimmune disorder. The population of cells may comprise one or more target effector cells, target B-cells, or a combination thereof. The target effector cells may be T-cells, NK cells, macrophages, or a combination thereof. The target effector cells may express cell surface molecules such as CD3, CD16, CD14, CD64, and combinations thereof. The target B-cells may express cell surface molecules such as CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38. CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1 , BCMA, SLAMF7 (CS-1), GPRC5D, and combinations thereof.

[000121] The first conjugate and the plurality of additional conjugates may be administered to the subject in an amount ranging from about 0.001 mg/kg to about 500 mg/kg, about 0.01 mg/kg to about 500 mg/kg, about 0.1 mg/kg to about 500 mg/kg, about 1 mg/kg to about 500 mg/kg, about 10 mg/kg to about 500 mg/kg, about 20 mg/kg to about 500 mg/kg, about 30 mg/kg to about 500 mg/kg, about 40 mg/kg to about 500 mg/kg, about 50 mg/kg to about 500 mg/kg, about 60 mg/kg to about 500 mg/kg, about 70 mg/kg to about 500 mg/kg, about 80 mg/kg to about 500 mg/kg, about 90 mg/kg to about 500 mg/kg, about 100 mg/kg to about 500 mg/kg, about 110 mg/kg to about 500 mg/kg, about 120 mg/kg to about 500 mg/kg, about 130 mg/kg to about 500 mg/kg, about 140 mg/kg to about 500 mg/kg, about 150 mg/kg to about 500 mg/kg, about 160 mg/kg to about 500 mg/kg, about 170 mg/kg to about 500 mg/kg, about 180 mg/kg to about 500 mg/kg, about 190 mg/kg to about 500 mg/kg, about 200 mg/kg to about 500 mg/kg, about 210 mg/kg to about 500 mg/kg, about 220 mg/kg to about 500 mg/kg, about 230 mg/kg to about 500 mg/kg, about 240 mg/kg to about 500 mg/kg, about 250 mg/kg to about 500 mg/kg, about 260 mg/kg to about 500 mg/kg, about 270 mg/kg to about 500 mg/kg, about 280 mg/kg to about 500 mg/kg, about 290 mg/kg to about 500 mg/kg, about 300 mg/kg to about 500 mg/kg, about 310 mg/kg to about 500 mg/kg, about 320 mg/kg to about 500 mg/kg, about 330 mg/kg to about 500 mg/kg, about 340 mg/kg to about 500 mg/kg, about 350 mg/kg to about 500 mg/kg, about 360 mg/kg to about 500 mg/kg, about 370 mg/kg to about 500 mg/kg, about 380 mg/kg to about 500 mg/kg, about 390 mg/kg to about 500 mg/kg, about 400 mg/kg to about 500 mg/kg, about 410 mg/kg to about 500 mg/kg, about 420 mg/kg to about 500 mg/kg, about 430 mg/kg to about 500 mg/kg, about 440 mg/kg to about 500 mg/kg, about 450 mg/kg to about 500 mg/kg, about 460 mg/kg to about 500 mg/kg, about 470 mg/kg to about 500 mg/kg, about 480 mg/kg to about 500 mg/kg, or about 490 mg/kg to about 500 mg/kg.

[000122] The ratio of the amount of first conjugate and the amount of the plurality of additional conjugates administered to the subject may range from about 1 ,000:1 to about 1 :1 ,000, about 900:1 to about 1 :1 ,000, about 800:1 to about 1 :1 ,000, about 700:1 to about 1 :1 ,000, about 600:1 to about 1 :1 ,000, about 500:1 to about 1:1,000, about 400:1 to about 1 :1 ,000, about 300:1 to about 1 :1,000, about 200:1 to about 1 :1 ,000, about 100:1 to about 1 :1 ,000, about 50:1 to about 1 :1 ,000, about 10:1 to about 1 :1 ,000, about 1,000:1 to about 1 :900, about 1 ,000:1 to about 1 :800, about 1 ,000:1 to about 1 :700, about 1 ,000:1 to about 1:600, about 1 ,000:1 to about

1 :500, about 1 ,000:1 to about 1 :400, about 1 ,000:1 to about 1:300, about 1 ,000:1 to about

1 :200, about 1 ,000:1 to about 1 :100, about 1 ,000:1 to about 1:50, or about 1,000:1 to about

1 :10.

[000123] The conjugates of the plurality of conjugates may be co-administered to the subject before the first conjugate is administered to the subject. The first conjugate may be administered to the subject from about 0.5 hours to about 72 hours, about 1 hours to about 72 hours, about 5 hours to about 72 hours, about 10 hours to about 72 hours, about 15 hours to about 72 hours, about 20 hours to about 72 hours, about 25 hours to about 72 hours, about 30 hours to about 72 hours, about 35 hours to about 72 hours, about 40 hours to about 72 hours, about 45 hours to about 72 hours, about 50 hours to about 72 hours, about 55 hours to about 72 hours, about 60 hours to about 72 hours, about 65 hours to about 72 hours, about 0.5 hours to about 65 hours, about 0.5 hours to about 60 hours, about 0.5 hours to about 55 hours, about 0.5 hours to about 50 hours, about 0.5 hours to about 45 hours, about 0.5 hours to about 40 hours, about 0.5 hours to about 35 hours, about 0.5 hours to about 30 hours, about 0.5 hours to about 25 hours, about 0.5 hours to about 20 hours, about 0.5 hours to about 15 hours, about 0.5 hours to about 10 hours, about 0.5 hours to about 5 hours, or about 0.5 hours to about 1 hours after the conjugates of the plurality of conjugates were administered to the subject.

[000124] The method can comprise repeating the contacting of the cells with the first conjugate. The method can comprise repeating the contacting of the cells with the plurality of conjugates. The method can comprise repeating the contacting of the cells with the first conjugate and the plurality of conjugates. The method can further comprise confirming apoptosis of the cells. Methods of confirming apoptosis are known to the art and may include but are not limited to: measuring caspase-3 activity, measuring annexin V / propidium iodine binding, measuring terminal deoxynucleotidyl transferase dUTP nick end-labeling, and combinations thereof. b. Methods of Treatment

[000125] Provided herein are methods of treating a subject in need thereof. The methods may include administering to a subject in need thereof a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; and administering to the subject in need thereof a plurality of additional conjugates. The plurality of additional conjugates may comprise a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of the target B-cell; and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell. The first morpholino oligonucleotide may hybridize with at least the second morpholino oligonucleotide, and the target effector cell induces apoptosis of the target B-cell, thereby treating the subject.

[000126] Administering may comprise intravenous administration. The method may comprise repeating the administration of the first conjugate. The method can comprise repeating the administration of the plurality of conjugates. The method can comprise repeating the administration of the first conjugate and repeating the administration of the plurality of conjugates.

[000127] The conjugates may target a population of cells in the subject. The subject may be a mammalian subject having cancer or an autoimmune disorder. The subject may have multiple myeloma, acute lymphocytic leukemia, Hodgkin’s lymphoma, non-Hodgkin's lymphoma, an organ transplant, rheumatoid arthritis, chronic lymphocytic leukemia, multiple sclerosis, systemic lupus erythematosus, autoimmune hemolytic anemia, pure red cell aplasia, idiopathic thrombocytopenic purpura, Evans syndrome, vasculitis, bullous skin disorders, type 1 diabetes mellitus, Sjogren's syndrome, Devic's disease, Graves' disease ophthalmopathy, or combinations thereof.

[000128] The population of cells may comprise one or more target effector cells, target B-cells, or a combination thereof. The target effector cells may be T-cells, NK cells, macrophages, or a combination thereof. The target effector cells may express cell surface molecules such as CD3, CD16, CD14, CD64, and combinations thereof. The target B-cells may express cell surface molecules such as CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38, CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1, BCMA, SLAMF7 (CS-1), GPRC5D, and combinations thereof.

[000129] The method can further comprise confirming apoptosis of the target B-cells in the subject. Methods of confirming apoptosis are known to the art and may include but are not limited to: measuring caspase-3 activity, measuring annexin V / propidium iodine binding, measuring terminal deoxynucleotidyl transferase dUTP nick end-labeling, and combinations thereof.

[000130] The first conjugate and the plurality of additional conjugates may be administered to the subject in an amount ranging from about 0.001 mg/kg to about 500 mg/kg, about 0.01 mg/kg to about 500 mg/kg, about 0.1 mg/kg to about 500 mg/kg, about 1 mg/kg to about 500 mg/kg, about 10 mg/kg to about 500 mg/kg, about 20 mg/kg to about 500 mg/kg, about 30 mg/kg to about 500 mg/kg, about 40 mg/kg to about 500 mg/kg, about 50 mg/kg to about 500 mg/kg, about 60 mg/kg to about 500 mg/kg, about 70 mg/kg to about 500 mg/kg, about 80 mg/kg to about 500 mg/kg, about 90 mg/kg to about 500 mg/kg, about 100 mg/kg to about 500 mg/kg, about 110 mg/kg to about 500 mg/kg, about 120 mg/kg to about 500 mg/kg, about 130 mg/kg to about 500 mg/kg, about 140 mg/kg to about 500 mg/kg, about 150 mg/kg to about 500 mg/kg, about 160 mg/kg to about 500 mg/kg, about 170 mg/kg to about 500 mg/kg, about 180 mg/kg to about 500 mg/kg, about 190 mg/kg to about 500 mg/kg, about 200 mg/kg to about 500 mg/kg, about 210 mg/kg to about 500 mg/kg, about 220 mg/kg to about 500 mg/kg, about 230 mg/kg to about 500 mg/kg, about 240 mg/kg to about 500 mg/kg, about 250 mg/kg to about 500 mg/kg, about 260 mg/kg to about 500 mg/kg, about 270 mg/kg to about 500 mg/kg, about 280 mg/kg to about 500 mg/kg, about 290 mg/kg to about 500 mg/kg, about 300 mg/kg to about 500 mg/kg, about 310 mg/kg to about 500 mg/kg, about 320 mg/kg to about 500 mg/kg, about 330 mg/kg to about 500 mg/kg, about 340 mg/kg to about 500 mg/kg, about 350 mg/kg to about 500 mg/kg, about 360 mg/kg to about 500 mg/kg, about 370 mg/kg to about 500 mg/kg, about 380 mg/kg to about 500 mg/kg, about 390 mg/kg to about 500 mg/kg, about 400 mg/kg to about 500 mg/kg, about 410 mg/kg to about 500 mg/kg, about 420 mg/kg to about 500 mg/kg, about 430 mg/kg to about 500 mg/kg, about 440 mg/kg to about 500 mg/kg, about 450 mg/kg to about 500 mg/kg, about 460 mg/kg to about 500 mg/kg, about 470 mg/kg to about 500 mg/kg, about 480 mg/kg to about 500 mg/kg, or about 490 mg/kg to about 500 mg/kg.

[000131] The ratio of the amount of first conjugate and the amount of the plurality of additional conjugates administered to the subject may range from about 1 ,000:1 to about 1 :1 ,000, about 900:1 to about 1 :1 ,000, about 800:1 to about 1 :1 ,000, about 700:1 to about 1 :1 ,000, about 600:1 to about 1 :1 ,000, about 500:1 to about 1:1,000, about 400:1 to about 1 :1 ,000, about 300:1 to about 1 :1,000, about 200:1 to about 1 :1 ,000, about 100:1 to about 1 :1 ,000, about 50:1 to about 1 :1 ,000, about 10:1 to about 1 :1 ,000, about 1,000:1 to about 1 :900, about 1 ,000:1 to about 1 :800, about 1 ,000:1 to about 1 :700, about 1 ,000:1 to about 1:600, about 1 ,000:1 to about

1 :500, about 1 ,000:1 to about 1 :400, about 1 ,000:1 to about 1:300, about 1 ,000:1 to about

1 :200, about 1 ,000:1 to about 1 :100, about 1 ,000:1 to about 1:50, or about 1,000:1 to about

1 :10.

[000132] The conjugates of the plurality of conjugates may be co-administered to the subject before the first conjugate is administered to the subject. The first conjugate may be administered to the subject from about 0.5 hours to about 72 hours, about 1 hours to about 72 hours, about 5 hours to about 72 hours, about 10 hours to about 72 hours, about 15 hours to about 72 hours, about 20 hours to about 72 hours, about 25 hours to about 72 hours, about 30 hours to about 72 hours, about 35 hours to about 72 hours, about 40 hours to about 72 hours, about 45 hours to about 72 hours, about 50 hours to about 72 hours, about 55 hours to about 72 hours, about 60 hours to about 72 hours, about 65 hours to about 72 hours, about 0.5 hours to about 65 hours, about 0.5 hours to about 60 hours, about 0.5 hours to about 55 hours, about 0.5 hours to about 50 hours, about 0.5 hours to about 45 hours, about 0.5 hours to about 40 hours, about 0.5 hours to about 35 hours, about 0.5 hours to about 30 hours, about 0.5 hours to about 25 hours, about 0.5 hours to about 20 hours, about 0.5 hours to about 15 hours, about 0.5 hours to about 10 hours, about 0.5 hours to about 5 hours, or about 0.5 hours to about 1 hours after the conjugates of the plurality of conjugates were administered to the subject.

7. Examples

[000133] The foregoing may be better understood by reference to the following examples, which are presented for purposes of illustration and are not intended to limit the scope of the invention. The present disclosure has multiple aspects and embodiments, illustrated by the appended non-limiting examples.

Example 1

Materials and Methods

[000134] Cell Lines, Antibodies, and Chemicals. The human cell lines Jurkat (Clone E6-1 ™), Raji (ATCC® CCL-86), MM.1S (CRL-2974™), HL-60 (CCL-240™), U266 (U266B1), Daudi (CCL-213™), RPMI 8226 (CCL 155™) were purchased from American Type Culture Collection (ATCC, Manassas, VA). Raji-Luc (CCL-86-Luc2™) cells were from Dr. Mark P. Chao, Stanford University. MM.1S-Luc, and KMS-12-BM cell lines were obtained from Huntsman Cancer Institute (Salt Lake City, UT). ANBL-6 cell line was obtained from Dr. Diane Jelinek (Mayo Clinic, Rochester, MN) and maintained in IMDM + 10% FBS with 1 ng/mL IL-6. The immortalized cell lines were cultured at recommended conditions in advised media from ATCC®. Jurkat, Raji, MM.1S, HL-60, U266, Daudi and RPMI 8226 were cultured in RPMI 1640 medium supplemented with 10% FBS, penicillin (200 U/mL) and streptomycin (200 pg/mL) at 37°C with 5% CO2. KMS-12-BM were cultured in RPMI 1640 supplemented as above with the addition of IL-6 (5 ng/mL) and 20% FBS. Human PBMCs were purchased from StemCell® (Vancouver, Canada; cat# 70025.3, lot# 211270301 C). T-Cells were isolated using negative selection and cultured in StemCell® (Vancouver Canada) T-cell media supplemented with CD3/CD28 mAb cocktail and recombinant IL-2 per manufacturer recommendations. T-cell activation was quantified through IFN-y release, CD69 expression, and doubling time. The monoclonal antibodies (mAb) used in MATCH conjugate synthesis or flow cytometric analysis were used as purchased.

[000135] Two complementary phosphorodiamidate morpholino oligonucleotide strands were customized and purchased from Gene Tools (Philomath, OR). The 25-base pair strands (SEQ ID NO: 1 and SEQ ID NO: 2) were modified with a primary amine at the 3’ termini (Chu et al., Theranostics 2015, 5(8): 834-846). Sequence: MORF1 5’- GAGTAAGCCAAGGAGAATCAATATA-NH2-3’ (SEQ ID NO: 53) and MORF2 5’- TATATTGATTCTCCTTGGCTTACTC-NH2-3’ (SEQ ID NO: 54). Tris(2-carboxyethyl) phosphine (TCEP), the heterobifunctional SM(PEG)2 linker, LysoTracker® green DND-26, JC-1 (5, 5’, 6,6’- tetrachloro-1 ,T3,3’-tetraethylbenzimidazoylcarbocyanine iodide), CCCP (carbonyl cyanide 3- chlorophenylhydrazone), PI (propidium iodide) and H2DCFDA (2’,7’-dichlorodihydrofluorescein diacetate) were purchased from Thermo Fisher Scientific® (Waltham, MA). Pepsin was purchased from Sigma-Aldrich® (St. Louis, MO). APC-annexin V was purchased from Biolegend® (San Diego, CA). Cy3- and Cy5-NHS (N-hydroxysuccinimide ester) were purchased from Lumiprobe (Cockeysville, MD). CCK-8 (Cell Counting Kit 8) was purchased from Dojindo (Kumamoto, Japan). TABLE 2 is a list of mAbs used for nanoconjugate synthesis and for immunostaining in vitro procedures.

TABLE 2. List of mAbs used for nanoconjugate synthesis and for immunostaining in vitro procedures outlined in the methods below.

[000136] Animals. All experiments involving animals were performed according to the protocol approved by the Institutional Animal Care and Use Committee (IACUC) and of the University of Utah. Female SCID C B-17 mice (Prkdc^/j were purchased from Charles River Laboratories (Wilmington, MA, USA) and used for xenograft models once body weight reached 17 g. Female

H2rg^,m1Wll/Sz (NRG) mice were purchased from The Jackson Laboratory (Bar Harbor, ME, USA) and used for xenograft models at 7 weeks.

[000137] Fab’-MORF Conjugate Synthesis and Characterization. The Fab’-MORF conjugates were synthesized as previously described (Summerton et al., Antisense Nucleic Acid Drug Dev. 1997, 7(3): 187-195; Chu et al., Theranostics 2015, 5(8): 834-846). Briefly, whole antibody was enzymatically cleaved using 10 w/w% pepsin in citric acid buffer pH 4.0 at 37 °C for 1.5 h. The reaction was monitored with FPLC (FIG. 3B and FIG. 3C). The F(ab’)₂ intermediate species was purified from the reaction mixture using centrifugal ultrafiltration (30 kDa MWCO filter 5* volume washes). F(ab’)₂ was reduced using TCEP (10 mM) in citric acid buffer pH 5.5 at 37 °C for 1 h. Free sulfhydryl groups were quantified with Ellman’s assay. Fab’ was retrieved using centrifugal ultrafiltration (10 kDa MWCO 5* volume washes). In parallel to the antibody workup, the morpholino oligonucleotide was conjugated to the bifunctional PEG linker through the 3’ primary amine and the NHS ester. MORF-NH₂ (1 eq.) was reacted with SM(PEG)2 (20 eq.) in a 1 :3 DMSO:PBS pH 7.4 mixture at r.t. for 2 h. The MORF-PEG2-maleimide product was purified using centrifugal ultrafiltration (3 kDa MWCO 8x volume washes). Freshly reduced Fab’ (1 eq.) and freshly prepared MORF-PEG2-maleimide (1.3 eq.) were reacted in PBS pH 6.5 for 3 h at ambient temperature. The final product was purified by centrifugal ultrafiltration (30 kDa MWCO 8x volume washes). MORF1 was conjugated to each B-cell Fab’ and MORF2 was conjugated to Fab’cD3.

[000138] Products were analyzed for purity using size exclusion chromatography (FIG. 5A). Hybridization between complementary conjugates was analyzed using dynamic light scattering (FIG. 5C), the hypochromic effect of base pair absorbance at 260 nm (FIG. 5B), and mass spectrometry (FIG. 4).

[000139] Cancer Cell Line Antigen Profiling. Immortalized cell lines including: i) B-cell lines of Raji (NHL), Daudi (NHL), MM.1S (MM), RPMI 8226 (MM), KMS-12-BM (MM), ANBL-6 (MM), U266 (MM); ii) the T cell line Jurkat (ALL); and iii) the myeloid line HL-60 (AML) were examined for their CD20, CD38, BCMA, and SLAMF7 expression levels. Cells were treated with fluorescently labeled primary antibodies and analyzed using cell sorting. Antigen expression was quantitated by normalizing geometric mean averages of labeled cells to untreated control cells.

[000140] MA TCH-lnduced T-Cell Activation and Cytotoxicity on Malignant Cell Lines. T arget cells (1 x 10⁵) were co-cultured with healthy donor, naive T-cells (1 x 10⁵) in a target cell-to-T cell ratio of 1 :1 for 2 h at 37 °C in a 24-well plate at a total volume of 400 pL RPMI 1640 cell culture medium. Cells were incubated with or without a premixed solution of Fab’_x-MORF1 and Fab’cD3-MORF2 at [MORF1] = [MORF2] = 50 nM. The dose was identified from IC₅o experiments of each single-target therapy with CCK-8 metabolic assay (Yajing et al., J. Clin. Oncol. 2020, 38(15 supplemental): 3034). IC50 values of each single-target therapy were between 500 pM and 10 nM. After 2 h incubation, cells were collected, washed with PBS, and stained with APC-annexin V and PI for 20 min at 4 °C. Percentage of target cells positive for both annexin V and PI were quantified using flow cytometry. Treatment groups were performed in triplicate and at least two replicates for each cell line were performed. Statistics were performed using one-way ANOVA and Tukey test.

[000141] Activating a Cohort of Naive T-Cells Against Multiple Cancers Using Cell-Specific MATCH. The same batch of naive, healthy donor T-cells were sequentially rechallenged with cancer cells over time. For the first challenge, T cells (2 x 10⁶) were added to a 6-well plate along with Raji B-cells (2 x 10⁵) in 4 ml_ RPMI 1640 cell culture medium. Cells were incubated with or without a premixed dose of Fab’cD2o-MORF1/Fab’cD3-MORF2 (50 nM) for 24 h at 37 °C. After 24 h, an aliquot (1 ml_) was removed and stained using APC-CD19 antibody. Number of CD19(+) cells in the untreated co-culture were qualitatively compared to the MATCH-treated cells. Remaining co-cultures were collected, washed with PBS, and resuspended in fresh RPMI 1640 culture medium. The second challenge commenced by adding MM.1S (1.5 x 10⁵) cells to each well and bringing the total volume to 4 ml_. Cells were incubated with or without a premixed dose of Fab’_BcMA-MORF1/Fab ’_CD3-MORF2 (50 nM) for 24 h at 37 °C. After 24 h, aliquot (1 mL) was removed and stained using PE-CD10 antibody. Number of CD10(+) cells in the untreated co-culture were compared to the MATCH-treated cells. Remaining co-cultures were collected, washed with PBS, and resuspended in fresh RPMI 1640 medium. The third challenge commenced with the addition of HL-60 (1.12 x 10⁵) cells to each well. Cells were incubated with or without a premixed dose of Fab’cD38-MORF1/Fab’cD3-MORF2 (50 nM) for 24 h at 37 °C. After 24 h, aliquots were stained with PE-CD33 antibody to compare number of HL-60 in untreated co-cultures to the MATCH-treated cells. The entire experiment was repeated using blinatumomab as the therapeutic agent dosed at 50 nM. All experiments were replicated two times.

[000142] MATCH-lnduced Mitochondrial Depolarization in Target Cancer Cells. Mitochondrial membrane integrity was tested using the mitochondrial membrane potential sensor JC-1 (Thermo Fisher Scientific (Waltham, MA)). T cell-to-target cell ratio of 1 :1 in a 48-well plate in 400 pL RPMI 1640 media were dosed with or without CD20-directed MATCH (50 nM) for 4 h at 37 °C. After 4 h, wells were collected, washed, and stained with JC-1 (4 pM) for 30 min at 37 °C. For positive depolarized control, untreated cells were treated with CCCP (0.5 pM) and incubated simultaneously with JC-1 for 30 min. After all JC-1 treatments, cells were washed analyzed using flow cytometry.

[000143] Quantifying the Effects of MATCH-Associated Caspase Activation. The attribution of caspase activation as a component of apoptosis in target cells was quantified by performing T- cell activation experiments as described above. Briefly, T cell-to-target cell ratio of 1 :1 in a 48- well plate in 400 pL RPMI 1640 media were dosed with or without CD20-directed MATCH (50 nM) for 4 h at 37 °C. After 4 h, wells were collected, washed, and stained with annexin V/PI for 20 min at 4 °C. Cells were again washed and analyzed for annexin V+/PI+ staining using flow cytometry. In this manner, target cells were first pre-treated with a pan caspase inhibitor, Z- VAD-FMK (5 pM), for 30 min at 37 °C. Cells were collected, washed, and used in the experimental conditions described. Pre-treated target cell apoptosis was compared to uninhibited target cell apoptosis levels to observe contribution of caspases to overall apoptosis.

[000144] MATCH-lnduced Reactive Oxygen Species in Target Cancer Cells. Levels of reactive oxygen species in the target cell cytosol after 4 h MATCH treatments were quantified by oxidation of 2’,7’-dichlorodihydrofluorescein diacetate (H₂DCFDA). Target cells were treated co-cultured in a T cell-to-target cell ratio of 1 :1 in a 48-well plate in 400 pL RPMI 1640 media were dosed with or without CD20-directed MATCH (50 nM) for 4 h at 37 °C. After treatment, cells were incubated with H₂DCFDA (5 pM) for 30 min at 37 °C. Cells were washed with PBS and analyzed using flow cytometry.

[000145] Dose Response of Multi-Targeted MATCH. The dose response of Multi-specific MATCH was assayed using a metabolic viability approach following a Chou-Talalay combination therapy setup (Gambles et al., J. Control. Rel. 2022, 350: 584-599; Chou et al., Cane. Res. 2010, 70(2): 440-446). Proper Chou-Talalay method requires prior knowledge of ECso values of individual therapies to construct a cocktail of combinational therapies at fold increases or fold decreases of the respective ECso values for each therapy in the cocktail. MM.1S cells were triple positive, thus, three single target dose response curves were attained for BCMA-, SLAMF7-, and CD38-directed MATCH, respectively. Briefly, a co-culture of naive, healthy donor T-cells with target myeloma cells (MM.1S) were plated in a 96-well plate in a 1:1 ratio (1x10⁴ of each cell per well). Cell co-cultures were dosed with serial dilutions of either BMCA-, SLAMF7-, or CD38-directed MATCH, from a dose range of 10 pM to 100 nM, for 24 h at 37 °C. After 24 h, CCK-8 (10 L per well) was added and incubated for 2 h at 37 °C. Absorbance at 460 nm was measured and plotted using GraphPad Prism using non-linear least squares fit. EC50 values for each single target therapy were attained and further used in the Chou-Talalay combination evaluation method. Briefly, serial dilutions above and below the respective EC50 values of each B-cell engaging Fab’-MORF1 were combined and dosed such that the final Fab’-MORF1 cocktail MORF1 concentration was combined with a one-to-one ratio of the complementary T-cell engager Fab’_CD3-MORF2 such that [MORF1]=[MORF2], For the multi-specific MATCH experiment, naive healthy T-cells were co-cultured in a 1 -to-1 ratio with MM.1S cells (1x10⁴ of each cell per well) in a 96-well plate. Premixed cocktails of Fab’scMA- MORF1, Fab’sLAMF7-MORF1, Fab’cD38-MORF1 , and Fab’cD3-MORF2 were titrated and added to each well. The cells were incubated for 24 h at 37 °C. After 24 h, CCK-8 (10 pL per well) was added and incubated for 2 h at 37 °C. Absorbance at 460 nm was measured and plotted using GraphPad Prism® using non-linear least squares fit.

[000146] MATCH-Treated Ex Vivo Chronic Lymphocytic Leukemia Patient Samples. Patient- derived Chronic Lymphocytic Leukemia (CLL) cells were obtained from the biobank of the Division of Hematology and Hematologic Malignancies at the Huntsman Cancer Institute. Every patient gave informed consent through a research protocol by the University of Utah Institutional Review Board (IRB# 45880). Primary blood mononuclear cells (PBMC)s were separated from whole blood using Ficoll-Paque centrifugation under sterile conditions. Cells were counted and seeded into a 96-well plate. The number of cells collected varied from patient-to-patient; thus, the number of cells loaded in each well differed for every patient. Opposed to 24-well plates, 96-well plates were used to facilitate cell-to-cell interactions. Each patient PBMC sample was stained with Cy5-daratumumab and Cy3-rituximab to quantify levels of CD38 and CD20 expression, respectively. Importantly, PBMCs were stained with Cy5-CD3 mAb and Fluor-421 CD24 mAb to determine the T cell-to-B cell ratio. PBMCs were dosed with single-target premixed MATCH solution targeting CD20 or CD38 (50 nM). In addition, separate PBMCs were dosed with a bispecific MATCH therapy where 25 nM Fab CD20-MORFI and 25 nM Fab’cD38- MORF1 were premixed with 50 nM Fab’_CD3-MORF2. PBMCs were treated with or without MATCH therapies for 3 h at 37 °C. After 3 h, cells were collected washed with PBS, and stained with APC-annexin V, propidium iodide, and Fluor-421 CD24 mAb. Target cells positive for annexin and propidium iodide were quantified using flow cytometry. Each patient sample experiment was performed in triplicate.

[000147] MATCH In Vivo Efficacy Using a Xenograft Human non-Hodgkin Lymphoma Murine Model. A small cohort, anti-cancer pilot study in vivo study consisting of 15 total mice distributed across five treatment groups (n=3 mice per group) was performed. Female C.B-17 SCID mice were inoculated via tail vein with either Raji B-cells (4x10⁵), or Raji B-cells (4x10⁵) and healthy donor, naive T-cells (4x10⁶) - administered as a co-culture mixture. Control cohorts consisted of Raji B-cells only, and Raji B-cells + T-cells only. Treatment groups consisted of MATCH (1 nanomole MORF1 equiv.; 60 pg), MATCH (0.01 nanomole MORF1 equiv.; 0.6 pg), and blinatumomab (0.6 pg, -0.01 nanomole) administered as a single dose tail vein injection 1 h post-inoculation. MATCH was administered in consecutive treatments of Fab’cD2o-MORF1 and Fab’cD3-MORF2 with a 5 h time lag between the two injections (Summerton et al., Antisense Nucleic Acid Drug Dev. 1997, 7(3): 187-195). The blinatumomab dose was injected at the time of Fab’cD3-MORF2 injection. The onset of hindlimb paralysis, indicating advanced disease onset, was monitored over time and mice were sacrificed upon late-stage disease onset (Yajing, J. Clin. Oncol. 2020, 38(15 supplemental): 3034; Scheetz et al., Nat. Biomed. Eng. 2019, 3(10): 768-782).

[000148] MATCH Optimization of T Cell-to-Target Cell Ratio In Vivo. Female NRG mice (n=5 mice per group) were inoculated via tail vein with luciferase expressing Raji B cells (4x10⁵) and various ratios of healthy donor, naive T-cells administered as co-culture mixtures. T cell-to-Raji cell ratios of 1 :1 (4x10⁵:4x10⁵), 2:1 (8x10⁵:4x10⁵), 3:1 (1.2x10⁶:4x10⁵), 5:1 (2x10⁶:4x10⁵), and 10:1 (4x10⁶:4x10⁵) were evaluated for maximum anti-cancer effect with a single 60 pg dose of CD20-directed MATCH. The control cohort consisted of Raji B-cells (4x10⁵) only. CD20- directed MATCH was administered as consecutive tail vein injections of Fab’_CD2o-MORF1 (1 nanomole MORF1 equiv.; 60 pg) 1 h post-inoculation, followed by Fab’_CD3-MORF2 (1 nanomole MORF2 equiv.; 60 pg) 6 h post-inoculation. The onset of disease was monitored with weekly I VIS imaging and development of hind-limb paralysis. Mice were sacrificed upon late-stage disease onset described by weight loss (> 20% body weight) and/or hind-limb paralysis. Example 2

Synthesis and Characterization of Fab’-MORF Conjugates

[000149] The ability to create a customized T-cell recruitment combination therapy comes from the generation of a malignant B-cell targeting motif library. B-cell antigen targets were strategically selected on several criteria: i) unique, or high prevalence of expression on malignant B-cells; ii) FDA approved monoclonal antibodies exist for the antigens; and iii) antigens spanning the three major hematological cancers including leukemia, lymphoma, and multiple myeloma. The four B-cell antigens chosen were CD20, CD38, B-Cell Maturation Antigen (BCMA), and SLAMF7 (FIG. 2).

[000150] Many bispecific antibodies (BsAbs) achieve T-cell activation by binding CD3 molecules in the TCR complex. Intracellular signaling domains of CD3 molecules produce a TCR-like activation response upon antibody binding. Many BsAb interactions at the interface of the T-cell and the target cell induce a mechanical force upon the T-cell cytoskeleton which triggers lytic granule mobilization and accumulation at the point of cell-to-cell contact. Release of the cytotoxic compounds from lytic granules kills the target cancer cell. Therefore, a T-cell engaging molecule based on an a-CD3 antibody was fashioned (FIG. 2).

[000151] MATCH conjugates were synthesized as previously described (Chu et al., Theranostics 2015, 5(8): 834-846; Gambles et al., Molecules 2021 , 26(15): 4658). Antibodies were first enzymatically digested with pepsin to remove the constant fragment (Fc) below the disulfide bonds in the hinge region (FIG. 3A). The disulfide bonds of the F(ab’)2 intermediate product were reduced to generate desired Fab’ products with reactive thiols on the truncated heavy chain. Complementary morpholino oligonucleotide strands (25 bp), functionalized with a primary amine at the 3’ end, were used as recognition motifs between the B-cell and T-cell conjugates. The oligonucleotides were conjugated to desired Fab’s through a bifunctional SM(PEG)2 linker. Primary amine groups of the oligonucleotides were reacted with the electrophilic ester and attached to the Fab’ molecules through a thiol-maleimide addition reaction. B-cell targeting Fab’ molecules were always conjugated with the identical oligonucleotide strand (MORF1); and T-cell targeting Fab’_CD3 molecules were always conjugated with the complementary oligonucleotide strand (MORF2) (FIGS. 3 and FIG. 4). TABLE 3 contains the MORF/Fab’ Ratios for FIG. 3C.

[000152] B-cell targeting motifs based on rituximab (Fab’_CD2o-MORF1), daratumumab (Fab’cD38-MORF1), a-BCMA (Fab’_BcMA-MORF1), and O-SLAMF7 (Fab’_SLAMF7-MORF1) were synthesized as described. The T-cell engaging molecule (Fab’cD3-MORF2) was synthesized as described using the complementary oligonucleotide strand. Target ratio of MORF/Fab’ was 1 .0. Therefore, one Fab’cD3-MORF2 molecule will hybridize, via Watson-Crick base pairing chemistry, with one Fab’-MORF1 molecule forming a stable heterodimer. The MORF substitution ratio was obtained by determining the Fab’ concentration using BCA protein assay and MORF concentration by UV-Vis absorbance at 260 nm. The ratio of [MORF] to [Fab’] was deemed the substitution ratio (FIGS. 3A-D). Mass spectrometry confirmed a single substitution product (FIG. 4).

[000153] The reactions were monitored with size-exclusion chromatography (FIG. 5A). Hybridization between complementary Fab’-MORF pairs was characterized using dynamic light scattering (FIG. 5C), hypochromic effect (FIG. 5B), and mass spectrometry (FIG. 4). Furthermore, fluorescently labeled Fab’-MORF conjugates were synthesized to observe cell-to- cell interactions under confocal microscopy (FIG. 7 Bottom Panel).

Example 3

B-Cell Antigen Expression Profiling and Cell Line-Specific T-Cell Activation

[000154] Numerous B-cell lines were cultured to provide a comprehensive montage of the three principal hematological cancers: leukemia, lymphoma, and myeloma. Immortalized cell lines tested include: i) B-cell lines of Raji (NHL), Daudi (NHL), MM.1S (MM), RPMI 8226 (MM), KMS-12-BM (MM), ANBL-6 (MM), and U266 (MM); ii) the T-cell line Jurkat (ALL); and iii) the myeloid line HL-60 (AML). The malignant B-cell lines were examined for their CD20, CD38, BCMA, and SLAMF7 antigen expression. Cells were treated with fluorescently labeled primary antibodies and analyzed using cell sorting. FIGS. 8A-C highlight three representative cell lines of major hematological cancers: HL-60 (leukemia), Raji (lymphoma), and MM.1S (myeloma). Each cell type had different expression profiles of the target antigens. HL-60 had high CD38 expression, and low-level expression of CD20; Raji had high CD20 and high CD38 expression; and MM.1S had high levels of CD38, SLAMF7, and BCMA expression. With the expression profiles in hand, each malignant cell line was co-cultured with healthy donor, naive T cells in a 1 :1 T-to-target cell ratio for 2 h at 37 °C with single target MATCH nanoconjugates. T-cell activation was achieved only if the target cell expressed the corresponding antigen. For example, Raji cells were killed by effector T-cells when treating with Fab’_CD38-MORF1 or Fab’cD2o-MORF1 , but not when treated with Fab’_BcMA-MORF1 or Fab’_SLAMF7-MORF1 targeting motifs. Levels of apoptosis in target B-cells was quantified using annexin V/PI staining and analyzed using flow cytometry for annexin(+)/PI(+) cells. The tabulated values for each cell exposed to each MATCH therapy are shown in FIGS. 8A-C.

[000155] To further demonstrate the modular capabilities of our two-component T-cell activating system, T-cell rechallenge experiments (FIGS. 9A-H) were structured wherein the same well of healthy donor, naive T-cells were exposed to three different cancer cell lines over a 72 h period. Every 24 h, a new cancer cell was added to the T-cells along with a dose of MATCH specific to each cancer. The same T-cells would be exposed to three different cancers and activated to kill each cancer using a cancer cell-specific targeting motif (FIGS. 9A-H). Upon each rechallenge, the co-culture was incubated with a dose of MATCH (50 nM) and a small aliquot of cells was removed to stain for presence of remaining target B-cells. The first round had T-cells (2 x 10⁶) co-cultured with Raji (lymphoma) cells (2 x 10⁵) representing a 10:1 T-to- target cell ratio. The co-culture was dosed with a premixture of Fab’_CD2o-MORF1 and Fab’_CD3- MORF2. After 24 h, a small aliquot of the cell suspension was removed and stained for CD19. Only residual Raji B-cells would stain positive for CD19. There were no detectable CD19(+) cells remaining in the co-culture. The T-cells were collected from the well, centrifuged and resuspended in fresh media before adding the second cancer cell rechallenge. The myeloma cell line, MM.1S, was added in a 10:1 T-to-target cell ratio. The co-culture was incubated for 24 h with a dose of premixed Fab’BMCA-MORF1 and Fab’cD3-MORF2. After 24 h, a small aliquot was removed and stained for CD10. Only remaining MM.1S cells would stain positive for CD10. There were no residual MM.1S cells detected in the MATCH-treated co-culture. The T-cells were again collected, centrifuged, and resuspended in fresh media before adding the third cancer - HL-60 (leukemia) - in a 10:1 T-to-target cell ratio. The co-culture was incubated for 24 h at 37 °C with a dose of premixed Fab’cD38-MORF1 and Fab’cD3-MORF2. After 24 h, a small aliquot was removed and stained for CD33. Only residual HL-60 cells would stain positive for CD33, but no remaining HL-60 cells were detected. In the described experiment, MATCH was used to activate the same group of a healthy donor, naive T-cells against three different blood cancers using three different targeting motifs. Furthermore, using the same rechallenge experiment protocol, MATCH was compared to blinatumomab. Blinatumomab was only able to ablate the CD19(+) Raji cells but was unable to activate T-cells against either MM.1S or HL-60 cell lines (FIGS. 9A-H).

Example 4

Investigating MATCH-lnduced T-Cell Cytotoxicity Mechanisms in Target Cells

[000156] MATCH functions the same as a BsAb - bridging the interface between the target cancer cell and the effector cytotoxic T-cell creating an immune synapse. Binding of Fab’cos- MORF2 to the TCR initiates a TCR-like activation of the T-cell, leading to polarization and degranulation of cytotoxic components in the direction of the interfacing target cell. Perforin opens pores in the target cell membrane kickstarting many apoptosis cascades simultaneously. Calcium influx initiates mitochondrial depolarization in the target cell leading to cytochrome C release and increased Bax expression. Cytotoxic molecules and enzymes released from the T- cell enter the target cell through perforin-created pores. Various peptidases cleave proteins and activate caspases. Altogether, cytotoxic death of target cell is rapid and occurs on the order of 1-6 h. Mitochondrial depolarization, caspase activity, and cytosolic reactive oxygen species were quantified within target cells after 1 h exposure to CD20- directed MATCH-induced T-cell activation (FIG. 10) against Raji cells. MATCH-treated, co-cultured Raji and T-cells showed significantly increased mitochondrial depolarization, caspase activity reliance, and cytosolic reactive oxygen species over untreated control co-cultures.

[000157] Lytic granules can be tracked by immunostaining and observed using microscopy (FIG. 10). Using LysoTracker® green DND-26, Jurkat T-cell granules were tracked in a coculture of Raji B-cells with and without CD20-directed MATCH. Jurkat cells were pretreated with the LysoTracker® before co-culture. Target Raji cells are first immunostained with anti- CD19 antibody, then co-cultured with the LysoTracker®-loaded Jurkat cells in a 1 :1 ratio. Cells were then either left untreated or CD20-directed MATCH was added to observe differences in granule polarization between treated and untreated co-cultures. Raji cells (red) and Jurkat cells (green) can be seen interacting with each other in both the untreated and treated cohorts. In the MATCH-treated cohort, T-to-target cell synapses are characterized by concentrated green fluorescence of the T-cell’s lytic granules localized at the cell-to-cel I interface. Untreated control co-culture has similar cell agglomeration, but very little granule polarization at the cell-to-cell interfaces. From these images, it can be concluded that MATCH polarizes T-cell lytic compartments to the point of target cell contact, thus initiating cytotoxicity.

[000158] Cell viability assays were conducted on the multiple myeloma cell line MM.1S. The MM.1S cell line was found to be triple positive (CD38(+)/BCMA(+)/SLAMF7(+)) for three target antigens. Multiple target antigens on the cell surface allowed us to investigate potency of monospecific MATCH therapies against a tri-specific combination therapy using the Chou- Talalay drug combination method. The metabolic assay, cell counting kit-8 (CCK8), was used to assess cell viability of cells treated with serial dilutions of MATCH monotherapies versus a Fab’cD38-MORF1/Fab’_BcMA-MORF1/Fab’sLAMF7-MORF1 combination therapy. Monospecific MATCH therapy ECso concentrations were found first by adding 1x10⁴ MM.1S and 1x10⁴ cultured, naive T-cells per well (1:1 ratio) to a 96-well plate. Cells were treated using a concentration gradient of MATCH starting with 100 nM and diluting down to 10 pM. Cells were incubated for 24 h at 37 °C, followed by the addition of CCK-8 assay buffer. Absorbance at 460 nm was measured. Because only the target cells in each well would be depleted, ICso curves between 100% and 50% viability were observed (T-cells remained viable). The curves between 100% and 50% viability were thus normalized and plotted using a non-linear least squares function in Prism (FIGS. 19A-C). Monospecific ECso values were found to be in the single-digit nanomolar-to-hundreds of picomolar range: 4.6 ± 0.3 nM, 7.7 ± 0.6 nM, and 0.760 ± 0.03 nM for BCMA-, SLAMF7-, and CD38-directed MATCH, respectively. For the tri-specific experiment, the EC₅O values for each monospecific therapy were used to create a combination therapy curve based on multiples of respective EC₅o values. Specifically, 4.6 nM Fab’_BcwiA-MORF1 , 7.7 nM Fab’si.AMF7-MORF1, 0.76 nM Fab’_CD38-MORF1 were added in combination, then diluted down by halves to 1/32-fold. Also, concentrations higher than the ECso values were created by doubling the concentrations up to 32-fold. The ECso value for the tri-specific combination therapy was found to be 0.35 ± 0.02 nM. Multi-specific T-cell recruitment enables more antigen engagement per target cell resulting in more TCR crosslinking and more efficient immune synapses per equivalent Fab’ dose.

Example 5 T Cell-to-Target Cell Ratio Impact Response in Ex Vivo Patient Chronic Lymphocytic Leukemia Samples

[000159] In vitro T-cell activation against malignant B-cell lines provided compelling proof-of- concept data; however, to further validate MATCH as an effective naive T-cell activator, the platform ex vivo on four Chronic Lymphocytic Leukemia (CLL) patient whole blood samples. CLL was chosen as the model disease due to the large variability of patient-to-patient peripheral tumor burden. Patient whole blood contains B CLL cells and autologous naive T-cells at various ratios. Upon isolation of the patient’s peripheral blood mononuclear cells (PBMCs) with Ficoll- Paque, MATCH’S ability to activate the patient’s own T-cells against their CLL cells was assessed. Patient CLL-containing PBMCs were first assayed for target antigen expression, CD20 and CD38, using fluorescently labeled primary antibodies. Second, the T-cell and B-cell counts were quantified and depicted as a T-to-B cell ratio. Patient’s T-cell counts remained relatively constant from patient-to-patient; however, the tumor burden, defined as the number of cancerous B-cells, varied widely. Of the four CLL patient samples tested, samples with T-to-B cell ratios close to 3:1 showed significant response to MATCH therapy. An example CLL patient sample (Patient 1) had a 3:1 T-to-B cell ratio. Patient 1 was CD20(+) and CD38(+) and showed a complete response after 24 h to both MATCH monotherapies, as well as, CD20/CD38 combination therapy. Patients 2 and 3 had very low T-to-B cell ratios of 1 :13, and 1:20, respectively, and did not respond to CD20- or CD38-directed MATCH therapies. Patient 4 had a T-to-B cell ratio of 1 :4 and was CD38(-). Likely due to the low T-cell count and lack of CD38 expression, Patient 4 had a partial response to CD20-directed MATCH and no response to CD38-directed MATCH. The small sample size limits our ability to draw decisive conclusions; however, the small collection of patient samples does lay bare an important facet of T-cell recruiting therapies - the need for an appropriate level of T cell-to-target cell ratio for an adequate therapeutic response to T-cell recruiting immunotherapies. Indeed, tumor burden is strongly correlated to patient response in hematological malignancies.

[000160] In vitro, T cell-to-target cell ratio and specific target cell antigen expression can be controlled to produce optimal T-cell activation. Ex vivo patient sample experimentation revealed the complexity of T-cell recruiting therapies being used as anti-cancer modalities. The large variance in total CLL cell counts between patients exposes a clinical limitation when treating hematological cancers with T-cell recruiting therapies. T-cell recruiting therapeutic effectiveness is limited by two major factors i) levels of target antigen expression on target cancer cells, and ii) total T-cell counts compared to tumor burden in the patient’s peripheral blood. Insufficient levels of target antigens and/or low T cell-to-tumor cell counts hampers T-cell recruiting approaches efficacy. Significant efforts have been made towards understanding the importance of effector- to-target cell ratio of T-cell engaging immunotherapies in vitro, but the data for T cell-to-target cell counts in vivo is scarce. Hipp et al. investigated the correlation of effector-to-target cell ratio of bone marrow-derived ex vivo myeloma samples to T cell ratio with their BCMA/CD3 BiTE and found higher ratios of T-cells correlated with more potent EC50S at all time points tested (Hipp et al., Leukemia 2017, 31 (8): 1743-1751). Rogala et al. provided a robust summary of published clinical trial data for blinatumomab but the number of T-cells at time of infusion was not reported for any of these clinical trials (Rogala et al., Expert. Onion. Biol. Then 2015, 15(6): 895-908). Often, patient response to blinatumomab is correlated to tumor burden, age, and number of prior therapies, but T-cell counts at time of infusion are not currently considered. In 2016, Chen et al. described a bispecific immunomodulatory biologic therapy and concluded the concentration of the tri-molecular synapse formed between the drug, the target cell, and the effector cell, not simply drug concentration, was responsible for efficacy (Chen et al., Clin. Pharm. Therap. 2016, 100(3): 232-241). The in vitro and ex vivo data supports the notion that efficacy is not strictly reliant on drug dose, but rather highly dependent on T cell-to-target cell ratio. To better understand the in vitro and ex vivo observations and the importance of T-cell counts in vivo, in vivo models were designed that provide a high level of control over T cell-to- target cell ratio.

Example 6

Single Dose of CD20-Directed MATCH Eliminated NHL Tumors In Vivo: Pilot Study

[000161] A pilot in vivo efficacy study was performed to survey CD20-directed MATCH efficacy and dosing schedules using a non-Hodgkin’s lymphoma (NHL) model. Considering the above discussion, Day 0 inoculation and dosing was performed to provide strict control over T cel l-to- target cell ratio in the murine peripheral blood. Conceptually, there are two modes of administration possible: i) a premixed, pre-hybridized bispecific molecule where self-assembly of the two components are allowed to occur before dosing; or ii) a two-step administration where the B-cell targeting Fab’-MORF1 can be dosed first followed by the T-cell engager Fab’cD3- MORF2 in sequence. Based on previous in vivo experiments conducted in the lab, it was hypothesized that a two-step dosing approach would result in significant anti-cancer efficacy. Female C.B-17 SCID mice were inoculated via tail vein with either Raji B-cells (4x10⁵) or Raji B- cells (4x10⁵) and healthy donor, naive T-cells (4x10⁶) administered as a co-culture mixture (1 :10 Raji-to-T Cell ratio) on Day 0. Treatment doses were given 1 h post-inoculation on Day 0 as a single dose tail vein injection (100 pL). Treatment groups consisted of MATCH (60 pg), MATCH at 100-fold dilution (0.6 pg), and blinatumomab at a dose equivalent to 100-fold diluted MATCH dose (0.6 pg). MATCH was administered in consecutive treatments of Fab’_CD2o-MORF1 and Fab’cD3-MORF2 with a 5 h time lag between the two injections (FIG. 25A). The 5 h time lag between MATCH doses is important because steady state plasma concentration of Fab’_CD2o- MORF1 occurs in roughly 4-5 h post-injection. Clearance of unbound Fab’_CD2o-MORF1 from the plasma will improve efficiency of Fab’_CD3-MORF2 motifs hybridizing with CD20-bound targeting motifs. The blinatumomab dose was injected at the time of Fab’_CD3-MORF2 injection. A total of 15 mice randomly distributed across five treatment groups (n=3 mice per group) was performed. The onset of hind-limb paralysis, indicating advanced disease onset, was monitored over time and mice were sacrificed upon late-stage disease onset.

[000162] Mice body weight and onset of hind-limb paralysis was monitored post-treatment. Untreated control mice (brown) each presented with hind-limb paralysis around Day 25 postinoculation (FIG. 25B). Blinatumomab treated (0.6 pg) mice (green) also showed no significant improvement of survival to untreated mice. Conversely, 0.6 pg MATCH treated mice (blue) showed a trending increase in survival over the blinatumomab treated cohort, although not significant (p=0.08). CD20-directed MATCH administered as a single 60 pg dose observed 3/3 mice surviving to the experiment endpoint of 75 days. Upon reaching Day 75, mice were sacrificed, their bone marrow harvested and stained for human B-cell markers. No residual disease was observed in the three surviving mice (FIG. 26A-D). The difference in efficacy between blinatumomab and MATCH could be due to differences in hydrodynamic radii leading to improved clearance parameters of the Fab’-MORF1 construct over the bispecific T-cell engager (BiTE) construct. Dynamic light scattering experiments revealed an R_H = 8.7 nm for Fab’cD2o- MORF1 (FIG. 5C) compared to the reported bispecific single-chain dibody radius of R_H - 2.7 nm. Furthermore, previously reported pharmacokinetic data of a l-labeled Fab’cD2o- MORF1 targeting motif revealed an elimination half-life ti/2,b = 5.1 h in mouse. The three-fold increase in RH observed in the Fab’-MORF species could be improving plasma concentration half-life over BiTE constructs leading to longer exposure times of target cancer cells with the therapy. The short half-life of blinatumomab, reported as 1.25 ± 0.63 h, is a limiting factor for the drug in the clinic where patients are given a 28-day continuous infusion. Indeed, efforts are being made to improve the dosing schedule and patient convenience of blinatumomab.

Example 7 Determining Optimal T Cell-to-Target Cell Ratio for CD20-Directed MATCH in NRG Mice

[000163] A second in vivo efficacy study was performed to test the hypothesis that a greater T cel l-to- target cell ratio will induce a higher occurrence of drug-T cell-target cell synapses leading to improved drug response. The response to single-dose MATCH in SCID mice was profound, thus, the interest was with CD20-directed MATCH response in more severely compromised murine models where onset of disease is more rampant. Female NRG mice were inoculated via tail vein with Raji B-luciferase cells (4x10⁵) and healthy donor, naive T-cells at varying ratios (administered as a co-culture mixture on Day 0). T cell-to-target cell ratios of 1 :1 , 2:1, 5:1 , and 10:1 were administered to different cohorts of randomly distributed mice. All mice received the same CD20-directed MATCH dose of 60 pg Fab’cD2o-MORF1 , given 1 h post-inoculation on Day 0 as a single dose tail vein injection (100 pL), followed by 60 pg Fab’_CD3-MORF2, given 6 h postinoculation on Day 0 as a single dose tail vein injection (100 pL), as described in FIG. 28A (Chu et al., Theranostics 2015, 5(8): 834-846). A total of 25 mice distributed across five treatment groups (n=5 mice per group) was performed. The onset of disease was monitored by weekly IVIS imaging and by the onset hindlimb paralysis.

[000164] The NRG mice accepted the xenograft at a vigorous rate. Significant tumor burden was observed in control mice in IVIS images taken on Day 7 post-inoculation. By Day 21 all control mice were sacrificed due to severe disease onset. All mice receiving a dose of CD20- directed MATCH significantly improved animal survival over control mice. Upon sacrifice, bone marrow analysis confirmed presence of Raji cells and residual T-cells in mice who received T- cell inoculations (FIG. 30). Interestingly, the observed survival data and IVIS imaging does not support the proposed hypothesis that more T-cells will ultimately correlate with improved response. The T cell-to-target cell ratio of 5:1 (2x10⁶ T cells co-inoculated with 4x10⁵ Raji cells) responded to a 60 pg dose of CD20-directed MATCH better than the 10:1 cohort (4x10⁶ T cells co-inoculated with 4x10⁵ Raji cells). The improved survival trend of the 5:1 cohort over the other cohorts suggests there exists a phenomenon of optimal T-cell counts when dosing T-cell recruiting therapies and discredits the notion that more T-cells correlates to improved response at a given drug dose. One can postulate the observed decrease in response at the highest T cell-to-cancer cell ratio is due to the abundant T-cell population acting as a Fab’_CD3-MORF2 sink where T-cell engaging motifs are effectively mopped up by the profuse number of available CD3 receptors. The effect results in inefficiencies and waste when forming the drug-T cell-target cell synapses necessary for recruitment and cytotoxicity. An intriguing follow-up experiment is planned that will quantify CD3 receptors per T-cell and target receptor number per target cell and investigate two-component MATCH dosing with a more stochiometric approach. Analyzing bispecific cell recruitment strategies with a stochiometric approach could edify the way the field of immunotherapeutics thinks about T-cell recruiting therapies.

Example 8

Discussion

[000165] Herein, a novel approach to bispecific T-cell engager therapies was described. Four antigen targeting motifs: Fab’_CD2o-MORF1 , Fab’_CD38-MORF1 , Fab’_BcMA-MORF1 , and Fab’_Si_AMF7- MORF1 and a complementary T-cell engaging motif, Fab’_CD3-MORF2, were developed to recruit and engage cytotoxic T cells with malignant B-cell cancers. MATCH was found to activated healthy, naive T-cells against leukemia, lymphoma, and myeloma in vitro based on cell-specific antigen targeting. T-cell killing of target cells was achieved by T-cell degranulation, followed by mitochondrial and caspase-dependent mechanisms within the target cell. A pilot study using a human non-Hodgkin’s lymphoma xenograft model in mouse revealed a single dose of CD20- directed MATCH absolved 3/3 mice of disease (up to 75 days). Surviving mouse bone marrow was clear of CD10(+) or CD19(+) human B-cells. A second murine model investigated the importance of T cell-to-target cell ratio, as high tumor burden observed in ex vivo CLL patient samples resulted in less response to MATCH therapy. The in vivo T-cell ratio experiment revealed an optimal T cell-to-target cell ratio of 5:1 at a 60 pg MATCH dose 1 h post-inoculation.

[000166] MATCH provides a modular “split antibody”-like approach where combinations of oligonucleotide-linked Fab’ motifs can be constructed with relative ease. The two-component system allows for construction of modular, bispecific constructs with high throughput compared to BiTEs, or other related recombinant strategies. A library of MORF1 targeting motifs can be constructed and used as an off-the-shelf product that can be tailored in a patient-specific manner when a new hematological cancer diagnosis arrives in the clinic. A simple flow cytometric assay can be used to aid in patient cancer cell-specific antigen selection and patientspecific MATCH formulation. Furthermore, the two-component technology enables fine tuning of T-cell activation by controlling the dose of the T-cell engaging motif, Fab’cD3-MORF2, independent of the targeting motif dosing. The combination of better targeting with control over T cell activation will have significant impact on patient prognoses and outcomes in the clinic. MATCH platform offers a new paradigm of personalized immunotherapy with potential advantages in both safety and efficacy. [000167] MATCH is currently under development to treat hematological B-cell malignancies; however, the technology is not limited to liquid tumors. MATCH is a platform technology that can be adapted for any cell-to-cell interfacing disease application, such as cancer (solid and liquid tumor) and autoimmunity. Current clinical immunotherapeutics against such indications are plagued with cases of relapse due to disappearance of primary cancer cell targets. The two-component approach allows for modulization and customizability that is not achievable with traditional bispecific T cell engaging immunotherapies. The platform capability of MATCH offers a method of both cancer-specific optimization and effector cell identity to achieve prolonged responses to treatment with more resistance to antigen-negative relapse. Personalized targeting will provide patients with better prognoses, while tunable effector cell activation will allow physicians more control over unwanted, and dangerous, effector cell side effects (exhaustion and cytokine release syndrome).

Example 9

Multi-Antigen T-Cell Hybridizers (MATCH)

[000168] Synthesis of MATCH Nanoconjugates. Conceptually, the Fc portion of the antibody has been removed, and the two binding domains apart from each other has been reduced, leaving just the Fab’ domains. From here, a dimerizing mechanism between differing Fab’s is needed. A recognition mechanism with high specificity, high binding energy, and high fidelity would be ideal. Complementary morpholino oligomer nucleotide strands work great for this application due to their rapid and specific hybridization to complementary strands. Additionally, the morpholino backbone chemistry is different to that of its DNA counterpart. A morpholine ring replaces the deoxyribose ring of traditional DNA monomers, thus changing phosphodiester linkages to phosphorodiamidate linkages which are not enzymatically cleavable in vivo and are neutral in charge. The resulting morpholine rings and phosphorodiamidate bonds render morpholino nucleotides more stable than DNA, but still capable of Watson-Crick base pair complementation.

[000169] Synthesis of the nanoconjugates can be followed by size exclusion chromatography (SEC). As mass is removed or added to intermediates, peak elution volumes will change. For example, FIG. 3B shows the pepsin cleavage of a-CD3 antibody. The product peak F(ab’)₂ begins to appear as a shoulder (green) at about 17 mL, but over time emerges as the major product after 3.5 h. FIG. 3C is an SEC spectrum overlay of Ab (blue), F(ab’)₂ (red), and Fab’ (green). FIG. 3D also shows a gel chromatography of the Fab’cos intermediate compared to whole antibody.

[000170] B-Cell Targeting Motif Library and Antigen Expression Profiling. The major advantage in this split antibody technology is in the ability to generate a large targeting motif library. Each B cell targeting Fab’ is conjugated with a single MORF1 strand. To date, this disclosure has four main B-cell antigen targeting Fab’-MORF1 compounds: Fab’_CD2o-MORF1 , Fab’cD38-MORF1, Fab’_BcMA-MORF1 , and Fab’_SLAwiF7-MORF1. As long as the T-cell targeting Fab’cD3 is conjugated with the complementary morpholino oligomer strand, MORF2, hybridization between cancer receptor-bound Fab’x-MORF1 and T-cell receptor-bound Fab’cos- MORF2 can occur.

[000171] A total of seven cell lines, representing the three hematological malignancies were profiled for expression of CD20, CD38, B Cell Maturation Antigen (BCMA), and SLAMF7. Two lymphoma lines (Raji and Daudi), one leukemia line (HL-60), and four multiple myeloma cell lines (MM.1S, KMS-12-BM, RPMI 8226, and U266) were surveyed. Antigen expression was measured using primary antibody immunostaining at 4° C for 30 min and flow cytometry.

[000172] T-Cell Activation Using MATCH Nanoconjugates. To begin the discussion on MATCH-induced T-cell activation, an important distinction between various forms of T-cells is necessary. For in vitro testing, three different T-cell models exist: i) naive, primary T-cells isolated directly from peripheral blood mononuclear cells (PBMCs); ii) CD3/CD28-stimulated primary T-cells undergoing cell culture (still antigen naive but proliferating); and iii) immortalized T-cell lines, such as the malignant T-cell line Jurkat (T-cell leukemia). Each model T-cell has their strengths and weakness for use in vitro and the cell type used will be identified during each experimental method. In general, immortalized cells are the easiest to work with due to their rapid doubling time and propensity to undergo mitosis without external stimulation or additives to the culture media. Large numbers of Jurkat cells can be acquired rapidly; whereas, culturing of primary T-cells requires external stimulation (CD3 & CD28 antibodies) and additives to the culture media (interleukins).

[000173] Initial binding and efficacy experiments were performed using Jurkat T-cells and Raji B-cells. The ratio of T-to-B cell is important to consider when designing experiments. One CD8+ T-cell has the capacity to kill multiple (> 10) B cells, but only under centrifugal conditions [4], In a typical in vitro CAR-T experiment, the ratio of T-to-B cells is < 1 - meaning more target cells than effector T-cells. Indeed, much of the ongoing work on MATCH in vitro utilizes cell ratio variance to examine phenomenon like T-cell exhaustion and cellular apoptotic mechanisms. There remains much to optimize in this area for in vitro modeling.

[000174] The most fundamental hypothesis to test was whether hybridization of nanoconjugates can occur at the cell-cell interface. One way to test the hypothesis is to use microscopy. A CD20-targeting Fab’-MORF1 was labeled with a Cy5 fluorophore and a CD3- targeting Fab’-MORF2 was labeled with a Cy3 fluorophore via peripheral lysine residues and an NHS ester coupling reaction. Confocal microscopy was employed to observe a co-culture of Jurkat T-cells with Raji B cells in a 1 :1 ratio. The B- and T-cells were clearly identifiable by their unique florescence spectrum. Two magnified images of B-cells (green) interacting with T-cells (red) are shown. At the interface of the B- and T-cells, a co-expression of red and green is observed leading to a yellow color (arrows). The yellow color indicates both Cy3 and Cy5 fluorophores are present, suggesting hybridization may be occurring. Light microscopy images of the co-culture with or without Fab’cD2o-MORF1/Fab’cD3-MORF2 MATCH for 1 h tells a compelling story. In the untreated control figure, the co-cultured cells appear to have few interactions with one another no notable cell clustering. In contrast, the MATCH treated coculture is highly clustered with a very high number of cell-cell contacts. Taken together, the confocal and the light microscopy images indicate B-to-T cells interactions are favorable in the presence of MATCH nanoconjugates.

Example 10

MATCH-lnduced Cytotoxicity of Malignant B-Cells In Vitro

[000175] Culturing Primary, Naive T-Cells Isolated from PBMCs. Jurkat T-cells are useful for imaging experiments, but they are themselves a cancer cell line. To properly model T-cell activation in vivo it is better to use naive, healthy T-cells to mimic the type of T-cells MATCH would encounter in the body. There are two options: i) to order PBMCs (or whole blood) from a supplier, perform a T-cell isolation and use the freshly isolated T-cells immediately; or ii) isolate T-cells from PBMCs, then culture the cells using chemical stimulation to induce T-cell expansion. Experiments have been performed with both fresh and cultured naive T-cells with no difference in observable efficacy. However, several important considerations have been discovered when using cultured naive T-cells, which will be discussed. [000176] Frozen PBMCs (50 million) were purchased from StemCell® (Vancouver, Canada), along with a negative selection T-cell isolation kit. Negative selection entails an antibody cocktail that binds to all mononuclear cells in the mixture except T-cells. Magnetic resin is then added to the PBMC/antibody cocktail suspension. The resin binds to the antibodies in the cell suspension. The suspension is placed into a magnet where the resin pulls the unwanted cells to the wall of the tube allowing the user to poor out the suspension which contains the desired T- cells and nothing else. PBMCs were stained with a CD3 antibody and compared to the isolated PBMC cell suspension (FIG. 5A). A pure T-cell suspension was observed in the isolated fraction (CD3(+)) while a bimodal CD3 histogram was observed in the PBMCs indicating the presence of CD3(-) cells.

[000177] Isolated T-cells were cultured in T-Cell Culture Media with CD3/CD28 activator and IL-2 supplementation. After four days, microscopy images were taken and compared to images taken at day 0 (FIG. 5B). Clusters of cells indicate proliferating T-cells (non-adherent). Additionally, expression of the leukocyte activation marker CD69 was analyzed and a significant increase in expression was observed in the four-day cultured T-cells (FIG. 5C). T-cells cultured in this manner will be referred to as pseudo-activated T-cells because they are technically naive but have been stimulated to proliferate and have activation markers on their surface.

[000178] MATCH-lnduced Apoptosis in Malignant B-Cells. Cultured, naive T-cells were the chosen T-cell format for efficacy experiments. The use of freshly isolated T-cells would probably serve as a better model; however, the practicality of continuously obtaining freshly isolated T-cells on demand was not feasible. Therefore, a co-culture of pseudo-activated T- cells (2 x 10⁵) and target B-cells (2 x 10⁵) in a 1:1 ratio were incubated with or without CD20-, CD38-, BCMA-, or SLAMF7-MATCH monospecific therapies for 2 h at 37° C in a 24-well plate. After treatment, cells were collected, washed with PBS, and stained with annexin V/propidium iodide (PI) for 20 min at 4 °C. After staining, cells were washed with PBS and analyzed with flow cytometry. Target B-cells positive for annexin V and PI were tabulated and designated as apoptotic.

[000179] Apoptosis was induced in all the target B-cell lines tested. B-cells only underwent apoptosis when exposed to Fab’-MORF1 moieties corresponding to antigens on the surface of the respective cells. No non-specific T-cell activation was observed that could possibly be generated by presence of Fab’_CD3-MORF2. Fab’_CD3-MORF2 alone is not enough to induce T- cell cytotoxicity. A corresponding B-cell Fab’-MORF1 must also be bound to the target cell surface. Additionally, a co-culture of T and target cell control (no treatment added) was performed to test for non-specific T-cell activation against foreign cells. Raji cells saw no nonspecific T-cell activation, but HL-60 and MM.1S did induce about 15% apoptosis non- specifically. It is hard to say what this apoptosis is from, but most likely, surface proteins on HL- 60 and MM.1S trigger some non-specific T-cell activation (maybe antigen activated). Even with a small amount of background cytotoxicity observed in some cell line co-cultures, the MATCH treated cells showed a significant increase in cytotoxicity. Extending the treatment time to 6 h does result in a higher level of apoptosis induction; however, beyond this time point relative apoptosis levels start to go down as target cells begin to die and get washed away during workup. Annexin V apoptosis experiments work best in the 2-4 h range.

Example 11

Multi-Specific MATCH for Cell Specific T-Cell Activation

[000180] Rechallenging T-Cells with Malignant B-Cells. Now that MATCH efficacy and potency was understood a little more, understanding the effects of MATCH on T-cell exhaustion was of interest. Specifically, there was interest in investigating how many times the same batch of T-cells be rechallenged with cancer cells before the T-cells lose their cytotoxic capacity. After some optimization, a protocol was devised to rechallenge T-cells with cancer cells multiple times over multiple days. There is currently work being done on developing these assays further and measuring key T-cell exhaustion markers over time; however, there is some compelling initial data to discuss.

[000181] First, an experiment was designed to rechallenge the same batch of T-cells with Raji B-cells (lymphoma) three times over a period of three days. The ratio of T-cells to Raji cells was adjusted so that complete ablation of Raji cells was observed after at least two rounds of rechallenge. A ratio of 10:1 T cells-to-Raji cells and 24 h of co-culture incubation with CD20- targeting MATCH (50 nM) was found to allow for T-cell killing of 99% Raji B-cells. For proof-of- concept, a rechallenge experiment was set up to begin with 2 x 10⁶ T-cells and 2 x 10⁵ Raji cells in a 6-well plate in 4 mL total volume. Wells for untreated control and MATCH treated cells were made. Fab’_RTx-MORF1 / Fab’_CD3-MORF2 (50 nM) was added to the treated well. The plate was incubated at 37 °C for 24 h. After 24 h, an aliquot (1 mL) was removed from both the treated and untreated wells. The remaining 3 mL of cell suspension was centrifuged and resuspended in fresh media. The cells were added to clean wells. Another round of Raji cells (1 .5 x 10⁵) was added to both wells and total media volume was leveled to 4 mL. Fab’RTx- M0RF1 1 Fab’cD3-MORF2 (50 nM) was added to the treated well once again. The aliquot was centrifuged, washed with PBS, then stained with CD19 (a pan B-cell marker). The aliquots were analyzed using flow cytometry for the presence of CD19(+) cells, indicating the presence of B- cells in the co-culture. Three rechallenge rounds were performed in this manner and found complete ablation of Raji B-cells in each round in the MATCH-treated well. The untreated control had an increasing ratio of Raji B-cells that made up the majority of cell suspension ratio by the end of the experiment.

[000182] A second rechallenge experiment was devised to demonstrate the versatility of the technology. The same protocol as above was used to activate the same batch of T-cells against multiple rounds of cancer cells. The difference here: instead of rechallenging with the same cancer cell line each rechallenge, T-cells were rechallenged with a different cancer cell line each time. At each rechallenge, T-cells were activated using different Fab’-MORF1 targeting motifs each time. A lymphoma cell line (Raji B cells) was used in the first round with CD20-targeted MATCH. No Raji cells were observed after 24 h treatment. Then, the the T-cells were challenged with a multiple myeloma cells line (MM.1S) using BCMA-targeted MATCH. No MM.1S cells were observed after 24 h treatment. Finally, the T-cells were challenged with a leukemia cells line (HL-60) using CD38-targeted MATCH. No HL-60 cells were observed after 24 h treatment. In this experiment, it was demonstrated that MATCH can activate the same naive T-cells against three different hematological cancers using three different antigen targets.

[000183] To add even more credence to the multi-specificity of MATCH-induced T-cell activation, the FDA approved, bispecific antibody blinatumomab was purchased to perform a similar rechallenge experiment as described. Blinatumomab is a bispecific T-cell engager (BiTE) that targets CD3 on T-cells and CD19 on B-cells. In a similar mechanism as MATCH, blinatumomab bridges cytotoxic T-cells with target cancer cells and induces T-cell killing of the target cell. However, if the target cell does not express CD19, blinatumomab cannot successfully recruit T-cells to kill the CD19(-) target cells. Therefore, a similar rechallenge experiment was devised where T-cells were exposed to a series of cancer cells over three days. Raji (CD19+) cells were first, followed by MM.1S (CD19-) cells, and finally HL-60 (CD19-) cells. As expected, blinatumomab was only able to activate T-cells against the first round of cancer, the CD19(+) Raji lymphoma cells. The second and third cancers were left unabated. [000184] The foregoing description of the specific aspects will so fully reveal the general nature of the invention that others can, by applying knowledge within the skill of the art, readily modify and/or adapt for various applications such specific aspects, without undue experimentation, without departing from the general concept of the present disclosure. Therefore, such adaptations and modifications are intended to be within the meaning and range of equivalents of the disclosed aspects, based on the teaching and guidance presented herein. It is to be understood that the phraseology or terminology herein is for the purpose of description and not of limitation, such that the terminology or phraseology of the present specification is to be interpreted by the skilled artisan in light of the teachings and guidance.

[000185] The breadth and scope of the present disclosure should not be limited by any of the above-described exemplary aspects, but should be defined only in accordance with the following claims and their equivalents.

[000186] All publications, patents, patent applications, and/or other documents cited in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication, patent, patent application, and/or other document were individually indicated to be incorporated by reference for all purposes.

[000187] For reasons of completeness, various aspects of the invention are set out in the following numbered clauses:

[000188] Clause 1 . A kit comprising: (i) a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; (ii) a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of a target B-cell; and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell.

[000189] Clause 2. The kit of clause 1 , wherein the plurality of additional conjugates further includes at least a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the fourth targeting moiety is adapted to bind to a fourth antigen that is on the surface of the target B-cell.

[000190] Clause 3. The kit of either clause 1 or clause 2, wherein the second morpholino oligonucleotide and the third morpholino oligonucleotide have the same nucleotide sequence.

[000191] Clause 4. The kit of clause 3, wherein the second and third morpholino oligonucleotides are both 95% complementary to the first morpholino oligonucleotide.

[000192] Clause 5. The kit of clause 3, wherein the second and third morpholino oligonucleotides are both 100% complementary to the first morpholino oligonucleotide.

[000193] Clause 6. The kit of any one of clauses 1-5, wherein the first targeting moiety is coupled to the first morpholino oligonucleotide by a first linker, the second targeting moiety is coupled to the second morpholino by a second linker and the third targeting moiety is coupled to the third morpholino oligonucleotide by a third linker.

[000194] Clause 7. The kit of clause 6, wherein the first linker, the second linker, and the third linker are each independently from about 10 A to about 100 A in length.

[000195] Clause 8. The kit of either clause 6 or clause 7, wherein one or more of the first linker, the second linker, and the third linker comprises polyethylene glycol (PEG) or succinimidyl-4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), and wherein each of the first linker, the second linker, and the third linker have either the same or a different molecular weight relative to one another.

[000196] Clause 9. The kit of any one of clauses 1-8, wherein the first, second and third morpholino oligonucleotides are each independently from about 10 nucleotides to about 30 nucleotides in length.

[000197] Clause 10. The kit of any one of clauses 1-9, wherein the first targeting moiety is a first antibody or a first Fab’ fragment of the first antibody, and wherein the first antigen is selected from the group consisting of CD3, CD16, CD14, and CD64.

[000198] Clause 11. The kit of clause 10, wherein the first antibody is selected from the group consisting of anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), anti- CD14 (clone: UCHM-1), and anti-CD64 (clone 10.1). [000199] Clause 12. The kit of clause 10, wherein the first antigen is CD3.

[000200] Clause 13. The kit of clause 12, wherein the first antibody is anti-CD3 (clone: UCHT- 1).

[000201] Clause 14. The kit of any one of clauses 1-13, wherein the second targeting moiety is a second antibody or a second Fab’ fragment of the second antibody and the third targeting moiety is a third antibody or a third Fab’ fragment of the third antibody, and wherein the second and third antigens are each independently selected from the group consisting of CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38, CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1, BCMA, SLAMF7 (CS-1), and GPRC5D.

[000202] Clause 15. The kit of clause 14, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, and Talquetamab.

[000203] Clause 16. The kit of clause 14, wherein the second and third antigens are each independently selected from the group consisting of CD3, CD19, CD20, CD38, CD52, BCMA, and SLAMF7 (CS-1).

[000204] Clause 17. The kit of clause 16, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , SJ25-C1, Rituximab, Obinutuzumab, Daratumumab, Isatuximab, Alemtuzumab, Belantamab, and Elotuzumab.

[000205] Clause 18. The kit of any one of clauses 1-17, wherein the second and third conjugates are co-formulated.

[000206] Clause 19. The kit of clause 18, wherein the second and third conjugates are further co-formulated with the first conjugate. [000207] Clause 20. The kit of either clause 18 or clause 19, wherein the plurality of additional conjugates further includes at least a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide; wherein the fourth targeting moiety is adapted to bind to a fourth antigen that is on the surface of the target B-cell; wherein the fourth conjugate is co-formulated with the second and third conjugates.

[000208] Clause 21. The kit of any one of clauses 1-19, wherein the nucleotide sequence of the first morpholino oligonucleotide is SEQ ID NO: 2 and the nucleotide sequence of the second and third morpholino oligonucleotides is SEQ ID NO: 1.

[000209] Clause 22. The kit of clause 19, wherein the nucleotide sequence of the fourth morpholino oligonucleotide is SEQ ID NO: 1.

[000210] Clause 23. A method of inducing apoptosis of a target B-cell, the method comprising: (i) contacting a population of cells with a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; and (ii) contacting the population of cells with a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of the target B-cell; a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell; and wherein the first morpholino oligonucleotide hybridizes with the second morpholino oligonucleotide, and the target effector cell induces apoptosis of the target B-cell.

[000211] Clause 24. The method of clause 23, wherein the population of cells are in a subject.

[000212] Clause 25. The method of clause 24, wherein the subject is a mammalian subject having cancer or an autoimmune disorder.

[000213] Clause 26. The method of any one of clauses 23-25, wherein the second morpholino oligonucleotide and the third morpholino oligonucleotide have the same nucleotide sequence. [000214] Clause 27. The method of any one of clauses 23-26, wherein the first targeting moiety is coupled to the first morpholino oligonucleotide by a first linker, the second targeting moiety is coupled to the second morpholino by a second linker and the third targeting moiety is coupled to the third morpholino oligonucleotide by a third linker.

[000215] Clause 28. The method of clause 27, wherein the first linker, the second linker and the third linker are each independently is from about 10 A to about 100 A in length.

[000216] Clause 29. The method of either clause 27 or clause 28, wherein one or more of the first linker, the second linker and the third linker comprises polyethylene glycol (PEG) or succinimidyl-4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), and wherein each of the first linker, the second linker and the third linker have either the same or a different molecular weight relative to one another.

[000217] Clause 30. The method of any one of clauses 23-29, wherein the first, second and third morpholino oligonucleotides are each independently from about 10 nucleotides to about 30 nucleotides in length.

[000218] Clause 31. The method of any one of clauses 23-30, wherein the first targeting moiety is a first antibody or a first Fab’ fragment of the first antibody, and wherein the first antigen is selected from the group consisting of CD3, CD16, CD14, and CD64.

[000219] Clause 32. The method of clause 31 , wherein the first antibody is selected from the group consisting of anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), anti-CD14 (clone: UCHM-1), and anti-CD64 (clone 10.1).

[000220] Clause 33. The method of clause 31 , wherein the first antigen is CD3.

[000221] Clause 34. The method of clause 33, wherein the first antibody is anti-CD3 (clone: UCHT-1).

[000222] Clause 35. The method of any one of clauses 23-34, wherein the second targeting moiety is a second antibody or a second Fab’ fragment of the second antibody and the third targeting moiety is a third antibody or a third Fab’ fragment of the third antibody, and wherein the second and third antigens are each independently selected from the group consisting of CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38. CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1 , PD-1 , BCMA, SLAMF7 (CS-1), and GPRC5D. [000223] Clause 36. The method of clause 35, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, and Talquetamab. .

[000224] Clause 37. The method of clause 35, wherein the second and third antigens are each independently selected from the group consisting of CD3, CD19, CD20, CD38, CD52, BCMA and SLAMF7 (CS-1).

[000225] Clause 38. The method of clause 37, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , SJ25-C1 , Rituximab, Obinutuzumab, Daratumumab, Isatuximab, Alemtuzumab, Belantamab and Elotuzumab.

[000226] Clause 39. The method of any one of clauses 23-38, wherein the nucleotide sequence of the first morpholino oligonucleotide is SEQ ID NO: 2 and the nucleotide sequence of the second and third morpholino oligonucleotides is SEQ ID NO: 1.

[000227] Clause 40. The method of clause 25, wherein the first conjugate and the plurality of additional conjugates are administered to the subject in an amount ranging from about 1 pg/kg to about 500 mg/kg.

[000228] Clause 41. The method of clause 40, wherein the ratio of the amount of first conjugate and the amount of the plurality of additional conjugates administered to the subject ranges from about 1,000:1 to about 1 :1 ,000.

[000229] Clause 42. The method of any one of clauses 25, 40 and 41 , wherein the second and third conjugates are co-administered to the subject before the first conjugate is administered to the subject, and wherein the first conjugate is administered to the subject from about 0.5 hours to about 72 hours after the second and third conjugates were administered to the subject.

SEQUENCES

SEQ ID NO: 1

M0RF1-a

GAGTAAGCCAAGGAGAATCAATATA

SEQ ID NO: 2

M0RF2-a

TATATTGATTCTCCTTGGCTTACTC

SEQ ID NO: 3

MORF1-b

GAAACCGCTATTTATTGGCTAAGAACAGATACGAATCATA

SEQ ID NO: 4

MORF2-b

TATGATTCGTATCTGTTCTTAGCCAATAAATAGCGGTTTC

SEQ ID NO: 5

M0RF1-C

GTAAACGCGACAAATGCCGATAATGCTTCGATAATAAT

SEQ ID NO: 6

M0RF2-C

ATTATTATCGAAGCATTATCGGCATTTGTCGCGTTTAC

SEQ ID NO: 7

M0RF1-d

GACAGAGTTCACTATGACAAACGATTTCACGAGTAATA

SEQ ID NO: 8

M0RF2-d

TATTACTCGTGAAATCGTTTGTCATAGTGAACTCTGTC

SEQ ID NO: 9

M0RF1-e

CCTGATACAGAAGTAGAAAGCAGTCACGCAATATA

SEQ ID NO: 10

MORF2-e

TATATTGCGTGACTGCTTTCTACTTCTGTATCAGG

SEQ ID NO: 11

MORF1-f

GAACAACGAGAGGTGCTCAATACAGATATCAATCA SEQ ID NO: 12

MORF2-f

TGATTGATATCTGTATTGAGCACCTCTCGTTGTTC

SEQ ID NO: 13

MORF1-g

AGTCATAGATAGACAGAATAGCCGGATAAACT

SEQ ID NO: 14

MORF2-g

AGTTTATCCGGCTATTCTGTCTATCTATGACT

SEQ ID NO: 15

MORF1-h

GATACAGAAGTAGAAAGCAGTCACGCAATATA

SEQ ID NO: 16

MORF2-h

TATATTGCGTGACTGCTTTCTACTTCTGTATC

SEQ ID NO: 17

M0RF1-i

GGCATAGATAACAGAATAGCCGGATAAACT

SEQ ID NO: 18

M0RF2-i

AGTTTATCCGGCTATTCTGTTATCTATGCC

SEQ ID NO: 19

MORF1-j

GACCAGTAGATAAGTGAACCAGATTGAACA

SEQ ID NO: 20

MORF2-j

TGTTCAATCTGGTTCACTTATCTACTGGTC

SEQ ID NO: 21

MORF1-k

GAGTACAGCCAGAGAGAGAATCAATATA

SEQ ID NO: 22

MORF2-k

TATATTGATTCTCTCTCTGGCTGTACTC

SEQ ID NO: 23

MORF1-I

GTGAACACGAAAGAGTGACGCAATAAAT

SEQ ID NO: 24

MORF2-I

ATTTATTGCGTCACTCTTTCGTGTTCAC SEQ ID NO: 25

M0RF1-m

GAACTAATGCAATAACTATCACGAATGCGGGTAACTTAAT

SEQ ID NO: 26

M0RF2-m

ATTAAGTTACCCGCATTCGTGATAGTTATTGCATTAGTTC

SEQ ID NO: 27

M0RF1-n

AGATGACGATAAAGACGCAAAGATT

SEQ ID NO: 28

M0RF2-n

AATCTTTGCGTCTTTATCGTCATCT

SEQ ID NO: 29

M0RF1-0

GGACCAAGTAAACAGGGATATAT

SEQ ID NO: 30

M0RF2-0

ATATATCCCTGTTTACTTGGTCC

SEQ ID NO: 31

MORF1-p

GCTGAAAACCAATATGAGAGTGA

SEQ ID NO: 32

MORF2-p

TCACTCTCATATTGGTTTTCAGC

SEQ ID NO: 33

MORF1-q

GATGAAGTACCGACAAGATA

SEQ ID NO: 34

MORF2-q

TATCTTGTCGGTACTTCATC

SEQ ID NO: 35

MORF1-r

GACAGGATGAATAACACAGT

SEQ ID NO: 36

MORF2-r

ACTGTGTTATTCATCCTGTC

SEQ ID NO: 37

MORF1-S GCAGCAAACGAAGTATAT

SEQ ID NO: 38

M0RF2-S

ATATACTTCGTTTGCTGC

SEQ ID NO: 39

MORF1-t

GTCATAACAGAACAGGTA

SEQ ID NO: 40

MORF2-t

TACCTGTTCTGTTATGAC

SEQ ID NO: 41

MORF1-U

TCAAGACAGAAGGAT

SEQ ID NO: 42

MORF2-U

ATCCTTCTGTCTTGA

SEQ ID NO: 43

MORF1-V

TAGCAACATAGGAAG

SEQ ID NO: 44

MORF2-V

CTTCCTATGTTGCTA

SEQ ID NO: 45

M0RF1-W

CAGAGAGCATAT

SEQ ID NO: 46

M0RF2-W

ATATGCTCTCTG

SEQ ID NO: 47

MORF1-X

CAAGAGGTACAT

SEQ ID NO: 48

MORF2-X

ATGTACCTCTTG

SEQ ID NO: 49

MORF1-y

AAGAGGTACA

SEQ ID NO: 50 MORF2-y TGTACCTCTT

SEQ ID NO: 51

M0RF1-Z

AAGGACAGTA

SEQ ID NO: 52

M0RF2-Z

TACTGTCCTT

SEQ ID NO: 53

M0RF1-a with primary amine at 3’ termini

GAGTAAGCCAAGGAGAATCAATATA-NH2

SEQ ID NO: 54

M0RF2-a primary amine at 3’ termini

TATATTGATTCTCCTTGGCTTACTC- N H₂

Claims

CLAIMS What is claimed is:

1. A kit comprising:

(i) a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell;

(ii) a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of a target 13- cell; and a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell.

2. The kit of claim 1 , wherein the plurality of additional conjugates further includes at least a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the fourth targeting moiety is adapted to bind to a fourth antigen that is on the surface of the target B-cell.

3. The kit of either claim 1 , wherein the second morpholino oligonucleotide and the third morpholino oligonucleotide have the same nucleotide sequence.

4. The kit of claim 3, wherein the second and third morpholino oligonucleotides are both 95% complementary to the first morpholino oligonucleotide.

5. The kit of claim 3, wherein the second and third morpholino oligonucleotides are both 100% complementary to the first morpholino oligonucleotide.

6. The kit of claim 1 , wherein the first targeting moiety is coupled to the first morpholino oligonucleotide by a first linker, the second targeting moiety is coupled to the second morpholino by a second linker and the third targeting moiety is coupled to the third morpholino oligonucleotide by a third linker.

7. The kit of claim 6, wherein the first linker, the second linker, and the third linker are each independently from about 10 A to about 100 A in length.

8. The kit of either claim 6, wherein one or more of the first linker, the second linker, and the third linker comprises polyethylene glycol (PEG) or succinimidyl-4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), and wherein each of the first linker, the second linker, and the third linker have either the same or a different molecular weight relative to one another.

9. The kit of claim 1 , wherein the first, second and third morpholino oligonucleotides are each independently from about 10 nucleotides to about 30 nucleotides in length.

10. The kit of claim 1 , wherein the first targeting moiety is a first antibody or a first Fab’ fragment of the first antibody, and wherein the first antigen is selected from the group consisting of CD3, CD16, CD14, and CD64.

11. The kit of claim 10, wherein the first antibody is selected from the group consisting of anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), anti-CD14 (clone: UCHM-1), and anti-CD64 (clone 10.1).

12. The kit of claim 10, wherein the first antigen is CD3.

13. The kit of claim 12, wherein the first antibody is anti-CD3 (clone: UCHT-1).

14. The kit of claim 1 , wherein the second targeting moiety is a second antibody or a second

Fab’ fragment of the second antibody and the third targeting moiety is a third antibody or a third

Fab’ fragment of the third antibody, and wherein the second and third antigens are each independently selected from the group consisting of CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38, CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1, PD-1, BCMA, SLAMF7 (CS-1), and GPRC5D.

15. The kit of claim 14, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, and Talquetamab.

16. The kit of claim 14, wherein the second and third antigens are each independently selected from the group consisting of CD3, CD19, CD20, CD38, CD52, BCMA, and SLAMF7 (CS-1).

17. The kit of claim 16, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , SJ25-C1 , Rituximab, Obinutuzumab, Daratumumab, Isatuximab, Alemtuzumab, Belantamab, and Elotuzumab.

18. The kit of claim 1 , wherein the second and third conjugates are co-formulated.

19. The kit of claim 18, wherein the second and third conjugates are further co-formulated with the first conjugate.

20. The kit of claim 18, wherein the plurality of additional conjugates further includes at least a fourth conjugate comprising a fourth targeting moiety and a fourth morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide; wherein the fourth targeting moiety is adapted to bind to a fourth antigen that is on the surface of the target B-cell; wherein the fourth conjugate is co-formulated with the second and third conjugates.

21. The kit of claim 1 , wherein the nucleotide sequence of the first morpholino oligonucleotide is SEQ ID NO: 2 and the nucleotide sequence of the second and third morpholino oligonucleotides is SEQ ID NO: 1.

22. The kit of claim 19, wherein the nucleotide sequence of the fourth morpholino oligonucleotide is SEQ ID NO: 1.

23. A method of inducing apoptosis of a target B-cell, the method comprising:

(i) contacting a population of cells with a first conjugate comprising a first targeting moiety and a first morpholino oligonucleotide, wherein the first targeting moiety is adapted to specifically bind to a first antigen on the surface of a target effector cell; and

(ii) contacting the population of cells with a plurality of additional conjugates, including: a second conjugate comprising a second targeting moiety and a second morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the second targeting moiety is adapted to bind to a second antigen that is on the surface of the target 13- cell; a third conjugate comprising a third targeting moiety and a third morpholino oligonucleotide that is at least 90% complementary to the first morpholino oligonucleotide, wherein the third targeting moiety is adapted to bind to a third antigen that is on the surface of the target B-cell; and wherein the first morpholino oligonucleotide hybridizes with the second morpholino oligonucleotide, and the target effector cell induces apoptosis of the target B-cell.

24. The method of claim 23, wherein the population of cells are in a subject.

25. The method of claim 24, wherein the subject is a mammalian subject having cancer or an autoimmune disorder.

26. The method of claim 23, wherein the second morpholino oligonucleotide and the third morpholino oligonucleotide have the same nucleotide sequence.

27. The method of claim 23, wherein the first targeting moiety is coupled to the first morpholino oligonucleotide by a first linker, the second targeting moiety is coupled to the second morpholino by a second linker and the third targeting moiety is coupled to the third morpholino oligonucleotide by a third linker.

28. The method of claim 27, wherein the first linker, the second linker and the third linker are each independently is from about 10 A to about 100 A in length.

29. The method of claim 27, wherein one or more of the first linker, the second linker and the third linker comprises polyethylene glycol (PEG) or succinimidyl-4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), and wherein each of the first linker, the second linker and the third linker have either the same or a different molecular weight relative to one another.

30. The method of claim 23, wherein the first, second and third morpholino oligonucleotides are each independently from about 10 nucleotides to about 30 nucleotides in length.

31. The method of claim 23, wherein the first targeting moiety is a first antibody or a first Fab’ fragment of the first antibody, and wherein the first antigen is selected from the group consisting of CD3, CD16, CD14, and CD64.

32. The method of claim 31, wherein the first antibody is selected from the group consisting of anti-CD3 (clone: UCHT-1), anti-CD16 (clone: 3G8), atibuclimab (anti-CD14), anti-CD14 (clone: UCHM-1), and anti-CD64 (clone 10.1).

33. The method of claim 31 , wherein the first antigen is CD3.

34. The method of claim 33, wherein the first antibody is anti-CD3 (clone: UCHT-1).

35. The method of claim 23, wherein the second targeting moiety is a second antibody or a second Fab’ fragment of the second antibody and the third targeting moiety is a third antibody or a third Fab’ fragment of the third antibody, and wherein the second and third antigens are each independently selected from the group consisting of CD5, CD19, CD20, CD22, CD25, CD30, CD33, CD37, CD38. CD40, CD47, CD52, CD70, CD74, CD79, CD80, CD123, PD-L1, PD-1, BCMA, SLAMF7 (CS-1), and GPRC5D.

36. The method of claim 35, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , Zolimomab, SJ25-C1 , Tafasitamab, Inebilizumab, Loncastuximab, Rituximab, Obinutuzumab, Ublituximab, Ocrelizumab, Ofatumumab, Ibritumomab, Epcoritamab, Glofitamab, Epratuzumab, Inotuzumab, Pinatuzumab, Camidanlumab, Inolimomab, Brentuximab, Iratumumab, Gemtuzumab, Lintuzumab, Vadastuximab, Lilotomab, Naratuximab, Otlertuzumab, Tetulomab, Daratumumab, Isatuximab, Elranatamab, Dacetuzumab, Bleselumab, Iscalimab, Lucatumumab, Ravagalimab, Selicrelumab, Teneliximab, Vanalimab, Lemzoparlimab, Magrolimab, Alemtuzumab, Vorsetuzumab, Milatuzumab, Polatuzumab, lladatuzumab, Galiximab, Talacotuzumab, Atezolizumab, Avelumab, Durvalumab, Cempilimab, RMP1-14, Dostarlimab, Retifanlimab, Toripalimab, Nivolumab, Pembrolizumaab, Belantamab, Elotuzumab, and Talquetamab. .

37. The method of claim 35, wherein the second and third antigens are each independently selected from the group consisting of CD3, CD19, CD20, CD38, CD52, BCMA and SLAMF7 (CS-1).

38. The method of claim 37, wherein the second and third antibodies are each independently selected from the group consisting of UCHT-1 , SJ25-C1, Rituximab, Obinutuzumab, Daratumumab, Isatuximab, Alemtuzumab, Belantamab and Elotuzumab.

39. The method of claim 23, wherein the nucleotide sequence of the first morpholino oligonucleotide is SEQ ID NO: 2 and the nucleotide sequence of the second and third morpholino oligonucleotides is SEQ ID NO: 1.

40. The method of claim 25, wherein the first conjugate and the plurality of additional conjugates are administered to the subject in an amount ranging from about 1 pg/kg to about 500 mg/kg.

41. The method of claim 40, wherein the ratio of the amount of first conjugate and the amount of the plurality of additional conjugates administered to the subject ranges from about 1 ,000:1 to about 1 :1 ,000.

42. The method of claim 25, wherein the second and third conjugates are co-administered to the subject before the first conjugate is administered to the subject, and wherein the first conjugate is administered to the subject from about 0.5 hours to about 72 hours after the second and third conjugates were administered to the subject.